DEAZAAMINOPTERINS FOR TREATMENT OF INFLAMMATION

Abstract: 5-Alkyl, 5-alkenyl, 5-alkynyl, and heteroaroyl-5-deazaaminopterins and 5,10-dideazaaminopterins are provided, as well as a method and composition employing such compounds for the treatment of inflammatory disease, such as rheumatoid arthritis.

Description: nEAZAAMTNOPTERTNS FOR TREATMENT OF TNFT.AMMATION Field of the Invention This invention relates to certain deazaaminopterin compounds useful for the treatment of inflammatory disease, such as rheumatoid arthritis, as well as a process for making such compounds and a method of using such compounds. Background of the Invention DeGraw et al., U. S. Patent 4,369,319, issued January 19, 1883, disclose a class of 10- deazaaminopterin compounds having the structure of formula: CH, COOH In the compound 10-deazaaminopterin, Ri and R2 are both hydrogen. In the alkyl derivatives of Patent No. 4,369,319, either or both of Ri and R2 is alkyl having from one to about eight, preferably one or two, carbon atoms. When only one of Ri and R2 is alkyl, the other is hydrogen. Exemplary Ri and R2 alkyl include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, amyl, iso-amyl, sec-amyl, tert-amyl, hexyl, iso-hexyl, heptyl, iso- heptyl, octyl, iso-octyl, 2-ethyl hexyl and tert-octyl. DeGraw et al., J.Med.Chem.. J , 552 (1974), report on the synthesis and antifolate activity of 10-deazaaminopterin. The antimicrobial and antitumor activities of the powerful dihydrofolic reductase inhibitors aminopterin and its N-10 methyl derivative, methotrexate, are well known, and numerous analogues have been made to further improve the potency, cell penetration, and toxicity properties of these compounds. As part of a continuing program to investigate structure-activity relationships in folic acid analogues, DeGraw et al. were interested in the effects of replacement of the nitrogen atom in the side chain of aminopterin and reported on the synthesis and biological activity of 10-deazaaminopterin. Continuing work with 10- deazaaminopterin and its 10-alkyl derivatives led to the discovery of their antileukemic activity, and to their efficacy in treating various ascites tumor systems. In accordance with U. S. Patent 4,369,319, it was determined that leukemia, as well as other malignancies, including ascitic tumors, can be ameliorated in warm-blooded lower animals by the administration of 10-deazaaminopterin, a nontrivial analogue of methotrexate, the current drug of choice for the treatment of leukemia in the clinic, as well as 10-alkyl derivatives of 10- deazaaminopterin, and it is expected that these compounds will have a similar effect in humans. Rheumatoid arthritis is an inflammation of the joints arising from infectious, metabolic, or constitutional causes, usually of unknown origin. It can result in serious restriction of movement and even invalidism. Since rheumatoid arthritis is a common disease that affects 2-3 million people in the United State alone, it poses a serious treatment problem. A substantial proportion of affected individuals will develop erosive joint disease and require surgical joint replacement despite therapies including disease-modifying antirheumatic drugs such as gold complexes, penicillamine, antimalarials, and methotrexate. In some patients with intractable rheumatoid arthritis, immunosuppressive agents including azathioprine, methotrexate, cyclophosphamϊde, and combinations of these drugs have proven beneficial. However, the potential side effects of some of these drags, including bone marrow toxicity and neoplasia, have limited their frequency of use and the dose that is given. The disease is one of a number of forms of proliferative disease, and the development of drugs for amelioration or curing the disease has occupied the attention of research organizations for many years, until most recently without appreciable success. The antifolic acid drug methotrexate has been used as an antitumor agent since 1955. Its cytotoxic action in tumors is related to its ability to inhibit (essentially irreversibly) the key enzyme, dihydrofolate reductase, required for biosynthesis of tetrahydrofolic acid. Tetrahydrofolate is a vital component in one-carbon metabolism in cells, being required for biosynthesis of purine and pyrimidine nucleosides of the DNA and RNA. The drug is a powerful cytotoxic agent whose principal toxicities occur with liver, kidney, and mucosal tissue, liver toxicity is the paramount concern for use in chronic therapy in a disease such as arthritis. The ability of methotrexate to affect the inflammatory conditions of rheumatoid arthritis may be linked to its cytotoxic behavior. This may be in the nature of immune suppression and could involve attack on inflammatory phagocytic cells such as macrophages or neutrophils and T-helper cells in the synovial region. Very few methotrexate analogs have been evaluated against arthritis in animals, and there is no clear indication whether the antiarthritic properties are directly proportional to cytotoxicity. Galivan et al.. Chem. Biol. Pteridines. DeGuyter. Berlin, 847 (1986), showed that adjuvant arthritis and streptoccocal cell wall arthritis in rats responded to doses of methotrexate relative to those used in man for treatment of rheumotoid arthritis. They also found that timing of dosage was most important for reduction of inflammation. Both methotrexate and aminopterin were found to inhibit inflammation, but other antifolate compounds that did not possess a 2,4-diaminopyrimidine unit or a benzoylglutamate side chain were ineffective. Piper et al., J. Med. Chem.. 25., 877-880 (1982), prepared the N-10 propyl, octyl, and propargyl analogues of methotrexate for evaluation. Biological evaluations of the three compounds consisted of studies of their effects on enzyme inhibition [(dihydrofolate reductase (EC 1.5.1.3) and thymidylate synthase)], L1210 cell growth inhibition, cellular membrane transport with various murine cell types (L1210, SI 80, Ehrlich, and epithelial), in vivo (mice) activity vs. L1210 leukemia and SI 80 ascites, and plasma clearance in mice. The in vivo results versus SI 80 ascites offered evidence that the propargyl compound might have a better therapeutic index against this tumor than methotrexate, but no other result from either of these compounds suggested significant superiority over methotrexate. Taylor et al., J. Org. Chem..48. 4852-4860 (1983) report that L-5-deazaaminopterin is equipotent with methotrexate both as an inhibitor of bovine liver dihydrofolate reductase and of L1210 murine leukemia cells. It is also equipotent with methotrexate in vivo both against L1210 and P388 leukemia in BDFi mice. Piper, J. Med. Chem.. 29_, 1080-1087 (1986) report evidence indicating that modifications at the 5-and 10-positions of classical folic acid antimetabolites lead to compounds with favorable differential membrane transport in tumor versus normal proliferative tissue. The 5-alkyl-5-deaza analogues were also investigated, including 5-methyl-5-deazaaminopterin, 5- methyl-5-deazamethotrexate, and 5-methyl-10-ethyl-5-deazaaminopterin. Biological evaluation of the 5-methyl-5-deaza analogues, together with previously reported 5-deazaaminopterin and 5- deazamethotrexate, for inhibition of dihydrofolate reductase (DHFR) isolated from L1210 cells and for their effect on cell growth inhibition, transport characteristics, and net accumulation of polyglutamate forms in L1210 cells revealed the analogues to have essentially the same properties as the appropriate parent compound, aminopterin or methotrexate, except that the last two were approximately 10 times more growth inhibitory than methotrexate. In in vivo tests against P388/O and P388/methotrexate leukemia in mice, the analogues showed activity comparable to that of methotrexate, with the more potent 20 producing the same response in the P388/O test as methotrexate but at one-fourth the dose; none showed activity against P388/methotrexate. DeGraw et al., J. Heterocyclic Chem.. £ 1 (9186), describe the synthesis of 5,10- dideazaaminopterin by two independent routes. Condensation of the piperidine enamine of 4-p- carbomethoxyphenylbutyraldehyde with ethoxymethylenemalononitrile followed by treatment of the resultant arylethylenaminomalononitrile with methanolic ammonia produced 2-amino-3- cyano-5-p-carbomethoxyphenethylpyridine. Cyclization of the aminocyanopyridine with • guanidine afforded 4-amino-4-deoxy-5, 10-dideazapteroic acid. Coupling of the pteroate intermediate with glutamate yielded the target 5,10-dideazaaminopterin. Alternatively, reduction of 2,4-diammo-6-formyl-5-deazapteιidine with sodium borohydride gave the 6- hydroxymethyl compound. Conversion to the bromide was followed by alkylation of dimethyl homoterephthalate to afford methyl 4-arnino-4-deoxy-10-carbomethoxy-5,10-dideazapteroate. Decarboxylation with ester cleavage (sodium cyanide in dimethyl sulfoxide at 180°) also gave the diaminopteroic acid. They reported that 5,10-dideazaaminopterin was an effective growth inhibitor of folate dependent bacteria, S.faecium and L. casei. Sirotnak et al, Cancer Research.5686-5691 (1988), describe studies examining a new class of 4-aminofolate analogues modified by an N to C conversion and alkyl substitution at the N-5 position of aminopterin and methotrexate. All of these analogues were equivalent to aminopterin and methotrexate as inhibitors of tumor cell dihydrofolate reductase (Ki = 3.49-5.16 pM). Piper et al., Chemistry and Biology of Pteridines. 1989, Walter de Gruyter & Co., Berlin - New York, state that modifications at the 5- and 10-positions of the classical antifolate structure have produced agents with antitumor activity superior to that of methotrexate. Examples are found in 5-alkyI-5-deaza analogues of aminopterin and methotrexate and in the lO-dea-aa inopterin series, particularly 10-ethyl-lO-deazaaminopterin whose ongoing clinical trials have produced highly favorable results). The 5-alkyl (methyl or ethyl) derivatives of 5- deazaaminopterin and 5-deazamethotrexate are equivalent to the parent compounds and to aminopterin and methotrexate as inhibitors of tumor cell dihydrofolate reductase, and overall, the activity of the 5-alkyl-5-deazamethotrexate derivatives appears to be equivalent to that of the 10-alkyl- 10-deazaaminopterin types. In continuing studies on the effects of changes at positions 5 and 10, they synthesized 10-ethyl-5-methyl-5,10-dideazaaminopterin, 10-propargyl-5- deazaaminopterin, and 10-propargyl-5-methyl-5-deazaaminopterin. The syntheses and available data from biological evaluations are reported. DeGraw et al., J. Med. Chem.. £2, 678 (1990), report the synthesis of the 10-methyl and 10- ethyl analogues of 5,10-dideazatetrahydrofolic acid (DDTHF), a potent inhibitor of glycinamide ribotide (GAR) formyltransferase. Key intermediates in the process were 10-methyl- and 10- ethyl-4-amino-4-deoxy-5,10-dideazapteroic acid. Condensation of the piperidine enamines of branched 4-(p-carbomethoxyphenyl)butyraldehydes with (acetoxymethylene)malo-nonitrile afforded l,l-dicyano-4-piperidinobutadiene. Subsequent reaction with alcoholic ammonium hydroxide yielded the appropriately substituted 2-amino-3-cyanopyridines. Ring closure with guanidine gave 10-methyl- and 10-ethyl-4-amino-4-deoxy-5,10-dideazapteroic acids. Coupling with diethyl glutamate followed by ester hydrolysis afforded 10-alkyl-5,10-dideazaaminopterin analogues which were effective inhibitors of DHFR derived from L1210, but were less potent than methotrexate for inhibition of growth of L1210 in culture. What is needed is an effective treatment for inflammatory disease, such as rheumatoid arthritis, which exhibits relatively low toxicity compared to current treatments. Description of the Invention In accordance with the present invention, 5-alkyl, 5-alkenyl, 5-alkynyl, and heteroaroyl- 5-deazaaminopterin and 5,10-dideazaaminopterin compounds are provided having the structure of Formula I: COOH wherein A is CH or N; X is one of and Ri is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms; and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms. Exemplary R and R2 alkyl include methyl, ethyl, propyl, iso-propyl, butyl, isobutyl, sec- butyl, tert-butyl, amyl, iso-amyl, sec-amyl, tert-amyl, hexyl, iso-hexyl, heptyl, iso-heptyl, octyl, iso-octyl, 2-ethyl hexyl andtert-octyl. Exemplary R2 alkenyl include allyl, 1-propenyl, crotyl (2-butenyl), 2-pentenyl, 4- pentenyl, 2-hexenyI, 5-hexenyl, 3-isopropenyl, 3-isobutenyl, and 4-octenyl. Exemplary R2 alkynyl include propargyl, 2-butynyl, 3-butynyl, 4-pentynyl, 5-hexynyl, and 7-octynyl. The invention also provides a process of treating rheumatoid arthritis and other proliferative diseases, which comprises administering to a warm-blooded animal having an inflammation of the joints or other evidence of the disease, a therapeutic nontoxic amount of a 5- alkyl, 5-alkenyl, 5-alkynyl, or heteroaroyl-5-deazaaminopterin or 5,10-dideazaaminopterin compound as defined by Formula I hereinabove, as such or in the form of a pharmaceutically acceptable salt thereof. These salts are formed with one or more free NH2 groups and/or COOH groups of the 5-deazaaminopterin or 5,10-dideazaaminopterin compound. Those compounds in which A is N, X is and Ri and R2 have from three to about eight carbon atoms, preferably from three to about five carbon atoms, are believed to be novel, and in addition, are exceptionally effective in the treatment of arthritis, in fact from six to ten times more effective than those compounds in which Ri and R2 have one or two carbon atoms, so as to constitute a delineated subclass within the larger genus. These compounds are therefore especially preferred. This subclass of compounds within the invention accordingly is defined by the structure of Formula II: COOH wherein Ri and R2 are alkyl, alkenyl, or alkynyl having from three to about eight, preferably from three to five, carbon atoms. Those compounds in which A is CH and X is and Ri and R2 are alkyl, alkenyl, or alkynyl having from three to about eight carbon atoms, preferably from three to about five carbon atoms, also are believed to be novel, and should be effective in the treatment of arthritis. This subclass of compounds within the invention accordingly is defined by the structure of Formula III: COOH wherein Ri and R2 are alkyl, alkenyl, or alkynyl having from three to about eight, preferably from three to five, carbon atoms. Exemplary 5-deazaaminopterin and 5,10-dideazaaminopterin compounds falling within Formulae I, II, or III are shown in the following Table IA. Table IA Compound No. Ri R2- 1 H H 2 H CH 3 3 H C 2 H 5 4 H C3H7 5 H CH 2 -=CHCH 2 - 6 H- CH≡CCH2- 7 H C5H11 8 H CgHπ CH3 H 10 CH3 CH3 11 CH3 C2H5 12 CH3 C3H7 13 CH3 CH 2 =CHCH 3 4 CH3 CH≡CCH2 5 CH3 C 8 Hi7 16 C 2 H 5 H 7 C 2 H 5 ___ CH3 8 C 2 H 5 C2H5 9 C 2 H 5 CH 2 =CHCH 2 Table IA Compound No. Ri R2 20 C2H5 CH≡≡CCH2 21 C3H7 H 22 C3H7 CH3 23 --C3H7 H 24 /-C3H7 CH3 25 n-C-tHg H 26 n-C4H9 CH3 27 CH 2 =CH-CH2- H 28 CH 2 =CHCH 2 CH 3 29 CH≡≡CCH 2 H 30 CH≡CCH 2 CH3 31 C5H11 H 32 CgHπ H One subclass of thienyl compounds and thienyl analogues within the invention is defined by Formula IN: CH, COOH wherein Ais N or CH; and Ri is hydrogen or alkyl having from one to about eight, preferably from one to three., carbon atoms; and R.2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight, preferably from one to three, carbon atoms. Another subclass of pyridyl compounds within the invention is defined by Formula V: CH 2 I COOH wherein A ϊs CH orN; and Ri is hydrogen or alkyl having from one to about eight, preferably from one to three, carbon atoms; and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight, preferably from one to three, carbon atoms. Exemplary deazaaminopterin compounds falling within Formulae I, IV, and V are shown in the following Tables IB and HB. Table IB 5-Peazaaminopterin Compounds (A = N Compound Rl R2 X No. Example A' 1 H H 2 H CH 3 3 H C2H5 Ok 4 H C3H7 O 5 H i-C 3 H 7 6 H C4H9 7 H C5H11 8 H C8H17 9 H CH2=CHCH2- 10 H HC≡CCH2- Example B' 11 CH 3 H Table IB 5-Deazaammopterin Compounds (A = MB Compound Rl R2 X No. Table IB 5-Deazaaminopterin Compounds (A = MB Compound Rl R2 X No. Table IB 5-Deazaaminopterin Compounds (A = N Compound Rl R2 X No. Table IB 5-Deazaaminopterin Compounds (A = f>B Compound Rl R2 X No. N- N c— Table IIB 5.10-Diazaaτninopterin Compounds (A = CBB Compound Rl R2 X No. Table IIB 5.10-Diazaaminopterin Compounds (A = CH) Compound Rl R2 X No. TaMe IB 5.10-Diazaaminopterin Compounds (A = CH) Compound Rl R2 X No. able ΠB 5.10-Diazaaminopterin Compounds (A = CH) Compound Rl R2 X No. The synthesis of the compounds of Formula π, wherein A is N and X is is adapted from that reported by Piper et al., J. Med. Chem.. 29, 1080-1087 (1986), as summarized in Procedure I below. RC(OC )3 + 2 Cli(CN)2 HCI 6a,b Procedure I The synthesis of compounds of Formulae I, IN, and N wherein A is Ν and X is any of the heterocyclic rings set out for these formulae can be carried out by Procedure -3, summarized hereinbelow. The synthesis of compounds of Formulae I, IN, and N wherein A is CH can be carried out by either of Procedures IE and IN, summarized hereinbelow. Procedures III and IN are believed to be novel and are the subject of co-pending patent applications. H,Ν CI X S„<^, COOH 11-1 II-5 II-4 II-6 II-7 Example F Procedure III III-1 Procedure IV The following Examples A-B represent typical preparations of compounds represented by Formulae I, E, and IE using Procedure I. Each compound was prepared at least twice by the procedure given. Reference numbers within the examples refer to steps or compounds prepared by steps as indicated in Procedure I or compounds listed in Table 1 A. Example A: Synthesis Of 5-Propvl-5-Deazaaτninopterin (6a) 2(a) 2-Amino-6-chloro-4-propyl-3,5-pyridinedicarbonitrile. A solution of trimethylorthobutyrate (la; 100 g, 0.670 mol), malononitrile (89.1 g, 1.35 mol), andpyridine (270 mL) was refluxed 1 hour. Excess pyridine was then removed by evaporation under reduced pressure (H2O aspirator, bath to 60°C). The residue was treated with 12 N HCl (1.15 L) and the mixture was transferred to a 5-L three-necked flask equipped with a thermometer, condenser, and mechanical stirrer (Teflon paddle). The mixture was stirred rapidly while being heated at 85-90°C for 1 hour. Solid material formed during this time. The mixture was cooled to 20- 25°C, and cold H2O (3 L) was added. After the mixture had been kept in a refrigerator overnight, the solid was collected, washed thoroughly with H2O and dried in vacuo. The product was homogeneous according to thin-layer chromatography (TLC) (EtOAc-cyclohexane, 1:1); yield 28% (42.4 g). Spectral data: mass, m/z 221, MH+ for C 1 0H9CIΝ4. 3(a) 2-Amino-4-propyl-3,5-pyridinedicarbonitrile. A solution of 2a (42.3 g, 0.193 mol) in dimetiiylformamide (DMF) (600 mL) and triethyl amine (Et3N) (70 mL) containing PdCtø (1.1 g) was shaken on a Parr apparatus under H2 at 45 psi (310.3 kPa) for 16 hours. Examination by TLC revealed conversion was incomplete. The mixture was filtered from catalyst with the aid of a little DMF. Fresh PdCl 2 (1.1 g) and more E. 3 N (35 mL) were added to the filtrate, and hydrogenation at 45 psi (310.3 kPa) was resumed. After 3 hours, TLC showed all 2a had been converted. The mixture was filtered and the filtrate was concentrated under reduced pressure (< 1 mm, bath 30°C) to about 75-100 mL. Dilution with cold H2O (1 L) caused precipitation of 3a; yield 91% (32.6 g), homogeneous by TLC. Spectral data: mas, m/z 187, MH + for C10H10N4; X H NMR (Me 2 SO-d6) d 0.95 (t, 3, CH3), 1.65 (m, 2, CH 2 ), 2.75 (t, 2, CH 2 ), 7.88 (br s, 2, NH 2 ), 8.52 (s, 1, C 6 -H). 4(a) 2,4-Diamino-5-propylpyrido[2,3-d]pyrimidine-6-carbonitrile. Anhydrous guanidine » HCl (6.15 g, 0.0640 mol) and NaOMe (3.49 g, 0.0650 mol) were combined in dry 2- (2-methoxyethoxy)ethanol (270 mL), and the mixture was stirred for about 0.5 hour before it was combined with a solution of 3a (12.0 g, 0.0640 mol) in 2-(2-methoxyethoxy)ethanol (335 mL). The stirred mixture was heated under N2 at 150-160°C for 7 hours. This mixture was allowed to cool to about 110°C while another solution of guanidine (one-half the previous amount) in 2-(2-methoxyethoxy)ethanol was prepared. The second guanidine solution was added, and heating at 150-160°C was resumed. After 5 hours, the. mixture was allowed to cool, then evaporated in vacuo (<1 mm) to a viscous mixture. Addition of cold H2O (~500 mL) gave a crude solid, which was collected and dried in vacuo. The crude product mixture (11.4 g) was dissolved in DMF, and the solution was swirled with silica gel (about 40 g of 60-200 mesh). Evaporation in vacuo as before gave a solid dispersion of crude product mixture and silica gel. The dispersion was pulverized, dried further in vacuo, then applied to a column (9 ¥ 50-cm) of silica gel (60-200 mesh poured from CHCI3). Gravity elution by CHCl3-MeOH (95:5) followed, and fractions homogeneous in 4a (R f -0.55 on TLC using CHCl3-MeOH, 5:1) were combined for evaporation to give pure 4a (3.3 g, 23% yield). Spectral data: mass, m/z 229, MH + for C 11 H12N6; *HNMR (Me2SO-dό) d 0.92 (t, 3, CH3), 1.62 (m, 2 CH 2 CH2CH3), 3.20 (m, 2, CH2CH 2 CH 3 ), 6.8-7.0 (br, 2, NH 2 ), 7.32 (br s, 2, NH ), 8.78 (s, 1, C 7 -H). 5(a) DiethylN-[4-[[(2,4-Dian-ino-5-propylpyrido[2,3-d]pyri-midin-6-yl)methyl] amino]- benzoyl]-L-glutamate. A stirred solution of 4a (1.21 g, 5.30 mmol) and diethyl N-(4- aminobenzoyl)-L-gIutamate (2.33 g, 7.23 mmol) in glacial AcOH (250 ml) containing damp Raney Ni (about 8 g) was kept under H2 at atmospheric pressure for approximately 4 hours or until H2 absorption from a gas buret had ceased near 240 ml. The catalyst was removed by filtration, and die filtrate was evaporated (H2O aspirator, bath 30°C). The residue was dissolved in die minimum volume of EtOH (12-15 mL), and die stirred solution was gradually treated witii 3% Na 2 CO 3 solution to pH 7.8. The resulting crude product mixture was collected with the aid of cold H2O, dried, and dispersed onto silica gel (60-200 mesh) as described above for precursor 4a. The dispersion was applied to a silica gel column (5 ¥ 50-cm) poured from CHCI3. Elution by gravity flow with CHCl3-MeOH (95:5) followed. After TLC showed all diethyl N-(4- aminobenzoyl)-L-glutamate and minor contaminants more mobile than 5a had been eluted, the system was switched to 85:15 CHCl3-MeOH. Fractions homogeneous with respect to 5a (R f -0.5 using CHCl3-MeOH, 3:1) were combined and evaporated to give pure 5a in 16% yield (470 mg). Spectral data: Mass, m/z 538, MH+; -Η. NMR d 0.92 (t, 3, CH3), 1.12-1.22 (21, 6, CH3CH2O overlapping), 1.54 (m, 2, CH 2 CH2CH3), 1.98 and 2.06 (2 m, 2, CHCH 2 CH 2 , nonequivalent), 2.42 (t, 2, CH2CH2CO), 3.02 (t, 2, CH2CH2CH3), 4.00-4.14 (brm, 4, CH 3 CH2O overlapping), 4.32 (d, 2, CH 2 NH), 4.38 (m, 1, CONHCH), 6.35 (br s, 2, NH 2 ), 6.56 (t, CH 2 NH), 6.66 and 7.68 (2 d, 4, C6H 4 ), 7.05 (s, 2, NH 2 ), 8.25 (d, 1, CONH), 8.52 (s, 1, C 7 - H). Anal. Calcd. for C27H 35 N7θ5« 0.5 H 2 O; C, 59.33; H, 6.64; N, 17.94. Found: C, 59.24, 59.55, H, 6.56, 6.49; N, 17.68, 17.71. 6(a) N-[4-[[(2,4-Dia-mno-5-propylpyrido[2,3-d]pyrirι-idin-6-yl)methyl]--mino]benzoyl3- L-glutamic Acid (5-Propyl-5-deazaaminopterin). A solution of 5a (400 mg, 0.732 mmol) in MeOH (800 mL) was treated with IN NaOH (1.7 ml), and the solution was kept at 20-25°C for 64 hours. MeOH was removed by evaporation under reduced pressure (H2O aspirator, bath 20- 25°C). Solution did not occur when the residue was treated witii H2O (50 ml), indicating that unchanged 6a was still present. The mixture was again made a clear solution by adding MeOH (300 ml) back to the aqueous suspension. The solution was left 48 hours longer at 20-25°C when high performance liquid chromatography (HPLC) [Piper et al., J. Med. Chem..29. 1080 (1986)] indicated that virtually all 5a had been converted to either 6a disodium salt or a monosodiu salt. MeOH was again removed, and the aqueous residue (adjusted to 50 ml) was treated with more IN NaOH (0.8 ml). After 4-5 more days at 20-25°C, HPLC showed that conversion to 6a was complete. The solution was clarified (Norit, Celite) and treated witii IN HCl to precipitate 6a at pH 3.8 as a pale beige solid; yield, 69% (260 mg), hydrated as indicated below. Assay by HPLC showed the product to be homogeneous. Spectral data: Mass, m/z 482, MH+; UN, lmax 228 nm (e 38,300), 299 (22,500) at pH 1; 225 nm (e 34,400), 284 (26,500) at pH 7; 225 nm (e 32,000), 284 (26,700) at pH 13; *H ΝMR (Me2SO-d6) d 0.92 (t, 3, CH 3 ), 1.56 (m, 2, CH2CH2CH3), 1.96 and 2.02 (two m, 2, CHCH2CH2, nonequivalent), 2.32 (t, 2 CH 2 CH 2 CO), 3.4 (t, 2, CH2CH2CH3), 4.30 (d, 2 CH 2 ΝH), 4.32 (m, 1, CONHCH), 6.54 (t, 1, CH 2 NH), 6.64 and 7.66 (2 d, 4, C6H4), 6.65 (br, 2 NH 2 ), 7.24 (br, 2, NH 2 ), 8.06 (d, 1, CONH), 8.52 (s, 1, C 7 - H). Anal. Calcd. for C23H 2 7N7O 5 «2 H 2 O: C, 53.38; H, 6.04; N, 18.94. Found: C, 53.72, 53.61; H, 5.86, 5.84; N, 18.74, 18.75. Example B: Synthesis Of 5-Butvl-5-DeazaaminoDterin 2(b) 2-Amino-6-chloro-4-butyl-3,5-pyridinedicarbonitrile. A solution of trimethyl orthovalerate (lb; 20.1 g, 0.124 mol) and malononitrile (16.4 g, 0.248 mol) in pyridine (50 mL) was refluxed 45 min, cooled, and evaporated. The residue was stirred with 12N HCl (210 mL) at 85°C (bath temp) for 45 min to give 2b as an insoluble solid. After dilution with H2O (100 mL), the mixture was chilled and the solid collected to give 2b in 27% yield (7.96 g); homogeneous by TLC (cyclohexane-EtOAc, 1:1). Spectral data: Mass, m/z 235, MH + for C11H11CIN4. 3(b) 2-Amino-4-butyl-3,5-pyridinedicarbonitrile. Hydrogenolysis of 2b (7.32 g, 31.2 mmol) in DMF (88 ml) containing PdCl2) and Et3N (9 ml) was conducted in a Parr shaker at 40 psi for 16 hours. Examination by TLC revealed absence of 2b. The catalyst was removed by filtration, and the filtrate was diluted with H2O to cause precipitation of 3b. The collected solid was reprecipitated from a Norit-treated and filtered (Celite) solution in DMF (120 ml) by adding H2O; yield, 90% (5.6 g); homogeneous by TLC (cyclohexane-ETOAc, 1:1). Spectral data: Mass, m/z 201, MH + for C11H12N4; ! H NMR (Me2SO-de) d 0.92 (t, 3, CH3), 1.38 (m, 2, CH2CH2CH2CH3), 1.58 (m, 2, CH2CH2CH2CH3), 2.75 (t, 2, CH2CH2CH2CH3), 7.88 (br, 2, NH 2 ), 8.52 (s, 1, C6-H). 4(b) 2,4-Dia-mno-5-butylpyrido[2,3-d]pyrimidine-6-carbonitrile. The annulation of 3b with guanidine was conducted as described for the conversion of 3a to 4a. A typical yield of pure 4b after column chromatographic purification (as described for 4a) was 20%; homogeneous by TLC (CHCl3-MeOH, 7:1). Spectral data: Mass, m/z 342, MH+ for Cι 2 Hι N 6 ; *H NMR d 0.88 (t, 3, CH3), 1.34 (m, 2, CH2CH2CH2CH3), 1.56 (m, 2, CH2CH2CH2CH3), 3.24 (t, 2, CH2CH2CH2CH3), 6.9 (br, 2, NH 2 ), 7.34 (br, 2, NH 2 ), 8.76 (s, 1, C 7 -H). .5(b) Diethyl N-[4-[[(2,4-Diamino-5-butylpyrido[2,3-d]pyrirnidin-6-yl)methyl]anιino]- benzoyl-L-glutamate. Reductive condensation of 4b with diethyl N-(4-aminobenzoyl)-L- glutamate was conducted as described for 5a. Pure 5b was also isolated as described for 5a. A typical yield of pure 5b was 15%. Spectral data: Mass, m/z 552, MH + for C28H37N7O5; ! H NMR (Me 2 SO-d6) d 0.85 (t, 3, CH3), 1.12-1.22 (21, 6, CH3CH2O, overlapping), 1.35 (m, 2, CH2CH2CH2CH3), 1.52 (m, 2, CH2CH2CH2CH3), 1.98 and 2.05 (2 m, 2, CHCH2CH2, nonequivalent), 2.42 (t, 2, CH 2 CH 2 CO), 3.04 (t, 2, CH2CH2CH2CH3), 4.0-4.15 (br m, 4, CH3CH2O, overlapping), 4.30 (d, 2, CH 2 NH), 4.38 (m, 1, CONHCH), 6.24 (br s, 2, NH 2 ), 6.52 (t, 1, CH 2 NH), 6.66 and 7.68 (2 d, 4, C5H4), 6.92 (br s, 2, NH 2 ), 8.24 (d, 1, CONH), 5.40 (s, 1, C 7 -H). 6(b) N-[[(2,4-Diammo-5-butylpyrido[2,3-d]pyrimidin-6-yl)methyl]-ιmino]benzoyl]-L- glutamic Acid (5-butyl-5-deazaaminopterin. A solution of 5b (50 mg, 0.091 mmol) in MeOH (150 ml) treated with IN NaOH (0.24 ml) was kept at 20-25°C for 4 days. MeOH was evaporated in vacuo (H2O aspirator, bath 25°C), and the residue was dissolved in H2O (3 mL). After 30 hours at 20-25°C, the solution was carefully treated with IN HCl to pH 3.8, where 6b precipitated; yield, 51% (24 mg). Assay by HPLC showed the product to be of 99.4% purity; spectral data: Mass, m/z 496, MH+; UN, l max 300 nm (e 23,900) atpH 1; 287 nm (e 25,900) at pH 1; 287 nm (e 26,100) at pH 13; *H ΝMR (Me2SO-d6) 0.85 (t, 3, CH ), 1.35 (m, 2, CH2CH2CH2CH3), 1.52 (m, 2, CH2CH2CH2CH3), 1.96 and 2.02 (2 m, 2, CHCH2CH3, nonequivalent), 2.32 (t, 2, CH 2 CH 2 CO), 3.06 (t, 2, CH2CH2CH2CH3), 4.3 (m, 3, CH 2 ΝH overlapping with CONHCH), 6.54 (t, 1, CH 2 NH), 6.66 and 7.66 (2 d, 4, C6H4), 7.18 (br s, 2, NH 2 ), 8.08 (d, 1, CONH), 8.54 (s, 1 C 7 H). Anal. Calcd. for C 2 4H29N 7 O 5 -1.5 H 2 O: C, 55.17; H, 6.17; N, 18.76. Found: C, 55.12; H, 6.03; N, 18.59. The synthesis of the compounds of Formula HI, wherein A is CH and wherein R2 is H, methyl or ethyl, is described by Taylor et al., J. Med. Chem.. 263. 914 (1985), and Taylor et al., U.S. Patent Nos. 4,845,216, issued My 4, 1989; and 4,684,653, issued August 4, 1987; and PTC International Publication No. O86105181 dated September 12, 1986. These references indicate these compounds have utility as antineoplastic agents, and so compounds wherein R2 is tiiree or more carbon atoms would be expected to have similar utility. The Examples A'-H 1 which follow illustrate application of Procedures E, El, and IV to die preparation of specific compounds of Formulae I, IV, and V, and represent preferred embodiments of the invention. Reference numbers within the examples refer to steps or compounds prepared by steps described in Procedures E, El, and IV or compounds listed in Table IB. Example A': Synthesis Of Compound 1. Table IB Bv Procedure II N-(5-Aminothiophene-2-carbonyl)-L-glutamic Acid Diethyl Ester (I-l). This compound was prepared by the method of Marsham et al., J. Med. Chem..24, 1594 (1991). 6-(Bromomethyl)-2,4-diaminopyrido[2,3-d]pyrimidine (1-3). This intermediate was prepared from 2,4-diaminopyrido[2,3-d]pyrimidine-6-methanol by the procedure of Piper et al., J. Med. Chem.. 35, 332 (1992). This particular preparation analyzed for a 1.75 HBr • 0.25 CH3COOH salt (formula wt. 406.7). The material was suitable for conversions to compounds I- 7, π-7, and m-7 in Table IB. N-[5-[[(2,4-Diaminopyrido[2,3-d]pyridin-6-yl)methyl]amino]dιiophene-2-carbonyl]-L- glutamic Acid (1-7). The bromomethyl compound 1-3 and the sidechain precursor I-l (1.2 mmol of each) were stirred with CaCO3 (2.4 mmol) in Me2NAc (15 mL) at 20-25°C for 4 days. The mixture was filtered, and die clear filtrate was added dropwise to excess 2.5% N-1HCO 3 solution witii stirring. The precipitate that formed was collected, dried, and chromatographed on silica gel with elution by CHCl3-MeOH (2:1) to give essentially pure diethyl ester; mass spectrum, m/e 502 (MH+ for C22H27N7O5S); predominantly one peak (>92%) by HPLC; yield 91 mg (15%). For hydrolysis, the ester (90 mg) was stirred with 1 N NaOH (1.8 mL) for 5 h (solution occurred after 1.5 h). Acidification to pH 3.8 caused precipitation of the product; yield 67 mg (75%) mass spectrum, m/e 446, MH+ for C 1 8H19N7O5S. Example B': Synthesis Of Compound 2, Table IB By Procedure II 6-(Bromomethyl)-2,4-diamino-5-methylpyrido[2,3-d]pyrimidine (1-4). 2,4- Diamino-5-methylpyrido[2,3-d]pyrimidine-6-methanol, [Piper et al., J. Med. Chem..3_ , 3002 (1992)] (4.5 g, 22.0 mmol) was dissolved in glacial AcOH (200 mL) at 95°C. The solution was cooled to 25°C, then treated with stirring with 30% dry HBr in AcOH (400 mL). When addition was complete, a clear solution remained. The flask was stoppered securely, and the solution was kept at 20-25°C before it was added to E-2O (2.2 L) with stirring. The precipitate that formed was collected under N2, washed with E-2O, and dried in vacuo (P2O5 and NaOH pellets); yield 9.5 g of 1-4 hydrobromide solvated by AcOH; yield 99% (based on formulation shown below), mass spectrum, m/e 268 and 270, MH+ for CgHioBrNs; 1H NMR (Me2SO-d6) d 2.78 (s, 3, CH3), 4.93 (s, 2, CH 2 Br), 8.17 (s, 2, NH2), 8.75 (s, 1, C7H), 9.32 (s, 2, NH 2 ); solvation by CH3CO2H evidenced by methyl-group singlet at d 1.90 whose integral height is one-half that of the CH3 group of 1-4. Thus the molar ratio of 1-4 to CH3CO2H is 1:0.5. Anal. Calcd for C9HioBrN 5 »1.7HBr«0.5CH3CO2H (formula wt. 435.7): C, 27.57; H, 3.17; N, 16.07. Found: C, 27.53, H, 3.37; N, 16.11. N-[5-[[(2,4-Diam o-5-memylpyrido[2,3-d]pvrimidin-6-yl)me yl]amino]thiophene-2- carbonyl]-L-glutamic Acid (1-8). Bromomethyl compound 1-4 (3.5 g, 8.0 mmol), sidechain precursor I-l (2.7 g, 8.2 mmol), and CaCO 3 (1.63 g, 16.3 mmol) were combined in Me2NAc (50 mL). The stiffed mixture was warmed briefly (5- 10 min.) at 70°C. Reactants other than CaCO3 dissolved readily. The mixture was stirred at 20-25°C in a stoppered flask under N2 for 7 days. Inorganic matter was filtered off, and silica gel (25 g) was added to the filtrate. The slurry was evaporated in vacuo, and the dispersion was pulverized for application on a silica gel column (4 x 50 cm). Elution by CHCl3-MeOH (95 : 5) removed front-running impurities. The product was eluted with CHCl3-MeOH (85 : 15). Appropriate fractions were combined and evaporated to give the diethyl ester of 1-8 (3.4 g, 81% yield). Mass spectrum, m/e 516, MH+ for C23H29N 7 O5S. 1-8 dietiiyl ester (258 mg, 0.50 mmol) was stirred 3 hours under N2 with IN NaOH (5.0 mL). The resulting clear solution was treated with 2N HCl to pH 3.5 to give 1-8; yield 46% (0.11 g). Mass spectrum, 460, MH+; 1HNMR (Me2SO-d6) d 1.88, 1.98 (two m, 2, CHCH2, nonequivalent), 2.30 (t, 2, CH2CO), 2.66 (s, 3, CH3), 4.25 (m, 1, CHCH2), 5.94 (d, 1, 4-ArH adjacent to 5-ArNH), 6.68 (s, 2, NH 2 ), 7.2-7.35 (m, 3, NH 2 overlapping 5-ArNH), 7.48 (d, 1, 3-ArH), 7.98 (d, 1, CONH), 8.52 (s, 1, C7-H). Anal. Calcdfor Cι 9 H2iN7θ5S»2.8H 2 O: C, 44.74; H, 5.26; N, 19.23. Found: C, 44.78; H, 5.18; N, 19.49. Example C: Synthesis Of Compound 12, Table IB, By Procedure II N-[5-(Methylaιι--ino)thiophene-2-carbonyl]-L-glutamic Acid Diethyl Ester (1-2). A solution containing I-l (1.78 g, 5.42 mmol), (i-Pr)2NEt (1.0 mL, 0.74 g, 5.7 mmol), and Me2SO4 (0.59 mL, 0.79 g, 6.2 mmol) in N,N-dimethylformamide (DMF, 20 mL) was warmed at 60°C for 2 h, then left at 20-25°C for 42 hours. The solution was evaporated in vacuo (1 mm, bath 25-30°C), and the residue was dissolved in EtOAc-cyclohexane (1: 1 by volume) for application to a silica gel column. Elution by the same solvent system afforded fractions homogeneous by TLC in 1-2. Fractions were combined and evaporated to afford 16% yield (287 mg) of 1-2 as an amber oil. Mass spectrum, m/e 343, MH+for C15H22N2O5S. N-[5-[[(2,4-Diaι----no-5-methylpyrido[2,3-d]pyriιnidin-6- yl)methyl]methyIammo]tMophene-2-carbonyl]-L-glutaιnic Acid (1-9). This compound was prepared from 1-4 (0.91 mmol) and sidechain precursor 1-2 (0.96 mmol) by essentially the same procedure as described above for the preparation of 1-8. After filtration, the reaction solution was evaporated (<1 mm, bath to 40°C). The residue was dissolved in CHCl3-MeOH (6 : 1) for application to a silica gel column. Elution by CHCl3-MeOH (6 : 1) gave fractions homogeneous by TLC (CHCl3-MeOH, : 1; Rf -0.5) which were combined and evaporated to give the diethyl ester of 1-9; yield 27% (132 mg), Mass spectrum, m/e 530, MH+ for C2 4 H3 1 N7O5S. This sample was hydrolyzed as described for 1-8 to give pure I-9»3H2θ in 80% yield (109 mg), Mass spectrum, m/e 474, MH+; 1H NMR (Me2SO-d6) d 1.90, 200. (two m, 2, CHCH 2 nonequivalent), 2.32 (t, 2, CH 2 CO), 2.60 (s, 3, 5-CH3), 2.87 (s, 3, CH3N), 4.30 (m, 1, CHCH 2 ), 4.50 (s, 2, CH 2 N), 6.05 (d, 1, 4-ArH adjacent to 5-ArN), 6.68 (s, 2, NH 2 ), 7.32 (s, 2, NH 2 ), 7.58 (d, 1, 3-ArH), 8.06 (d, 1, CONH), 8.35 (s, 1, C7H). Anal. Calcd for C 2θ H 23 N 7 O 5 S » 3H2θ: C, 45.53; H, 5.54; N, 18.59. Found: C, 45.60; H, 5.28; N, 18.36. Example D': Synthesis Of Compound 19, Table IB, By Procedure II 6-(Bromomethyl)-2,4-diamino-5-ethylpyrido[2,3-d]pyrimidine (1-5). 2,4- Diamino-5-ethylpyrido[2,3-d]pyrimidine-6-methanol (2.80 g, 12.8 mmol), prepared as described • for the 5-CH3 homolog [Piper et al., J. Med. Chem.. 3J>, 3002 (1992)], was converted to 1-5 as described above for 1-4; yield 5.3 g, Mass spectrum, m/e 282 and 284, MH+ for CιoHi2BrN5; 1H NMR (MeSO-d6) d 1.24 (t, 3, CH 3 ), 3.24 (q, 2, CH 2 ), 4.94 (s, 2, CH 2 Br), 8.14 (s, 2, NH ), 8.80 (s, 1, C7-H), 9.22 (s, 2, NH2); solvation by CH3CO 2 H is evidenced by methyl-group singlet at d 1.92 whose integral height is one-third that of the CH3 group of 1-5. Based on the results, the formulation for the product would be I-5»1.4HBr»0.33CH3Cθ2H (formula wt. 416.7). N-[5-[[(2,4-Diamino-5-ethylpyrido[2,3-d]-pyrimidin-6-yl)methyl]meuιylamino]- thiophene-2-carbonyl]-L-glutamic Acid (1-10). Alkylation of 1-2 by 1-5 (0.84 mmol of each) was done as described under the preparations of 1-8 and 1-9. Purification of the ester produced was done as described for 1-9 with CHCl3-MeOH (7 : 1) used as eluant. The yield of pure diethyl ester of 1-10 was 27% (124 mg); mass spectrum, m/e 544, MH+ for C25H33N7O5S. This sample was hydrolyzed as described for 1-8 (or 1-9) to give I-10»3H2O in 82% yield (102 mg), Mass spectrum, m/e MH+; 1H NMR (Me2SO-d6) d 1.16 (t, CH3CH2), 1.90, 2.00 (two m, 2, CHCH2 nonequivalent), 2.30 (t, 2, CH 2 CO), 2.86 (s, 3, CH3N), 3.00 (q, 2, CH3CH2). 4.28 (m, CHCH 2 ), 4.52 (s, 2, CH N), 6.05 (d, 1, 4-ArH adjacent to 5-ArN), 6.60 (s, 2, NH 2 ), 7.22 (s, 2, NH2), 7.61 (d, 1, 3-ArH), 8.06 (d, 1, CONH), 8.40 (s, 1, C7H). Anal. Calcd for C21H25N7O5S.-3H2O: C, 46.57; H, 5.77; N, 18.10. Found: C, 46.55,; H, 5.52; N, 18.15. Example E': Synthesis Of Compound 21, Table IB, By Procedure II Preparation of 6-Bromomethyl-2,4-diamino-5-propylpyrido(2,3-d)pyrimidine (I- 6). 2-Amino-6-chloro-4-propyl-3,5-pyridinedicarbonitrile. A solution of trimethyl orthobutyrate (la; 100 g, 0.670 mol), malononitrile (89.1 g, 1.35 mol), and pyridine (270 mL) was refluxed 1 hour. Excess pyridine was then removed by evaporation under reduced pressure (H2O aspirator, bath to 60°C). The residue was treated with 12 N HCl (1.15 L) and die mixture was transferred to a 5-L three-necked flask equipped with a thermometer, condenser, and mechanical stirrer (Teflon paddle). The mixture was stirred rapidly while being heated at 85-90°C for 1 hour. Solid material formed during this time. The mixture was cooled to 20-25°C, and cold H2O (3 L) was added. After the mixture had been kept in a refrigerator overnight, die solid was collected, washed tiioroughly with H 2 O and dried in vacuo. The product was homogeneous according to TLC (EtOAc-cyclohexane, 1:1); yield 28% (42.4 g). Spectral data: mass, m/z 221, MH+ for C10H9CIN4. 2-Amino-4-propyl-3,5-pyridinedicarbonitrile. A solution of 2a (42.3 g, 0.193 mol) dimediylformamide (DMF, 600 mL) and triethyl amine C&gN, 70 mL) containing PdCl2 (1.1 g) was shaken on a Parr apparatus under H2 at 45 psi (310.3 kPa) for 16 hours. Examination by TLC revealed conversion was incomplete. The mixture was filtered from catalyst with the aid of a little dimethyl formamide (DMF). Fresh PdCl 2 (1.1 g) and more Et3N (35 mL) were added to the filtrate, and hydrogenation at 45 psi (310.3 kPa) was resumed. After 3 hours, TLC showed all 2a had been converted. The mixture was filtered and die filtrate was concentrated under reduced pressure (< 1 mm, bath 30°C) to about 75-100 mL. Dilution with cold H2O (1 L) caused precipitation of 3a; yield 91% (32.6 g), homogeneous by TLC. Spectral data: mas, m/z 187, MH+ for C10H10N4; IH NMR (Me2SO-d6) d 0.95 (t, 3, CH3), 1.65 (m, 2, CH 2 ), 2.75 (t, 2, CH 2 ), 7.88 (br s, 2, NH 2 ), 8.52 (s, 1, Cβ-H). . 2,4-Diammo-5-propylpvrido[2,3-d]pyrimidine-6-carbonitrile. Anhydrous guanidine»HCl (6.15 g, 0.0640 mol) and NaOMe (3.49 g, 0.0650 mol) were combined in dry 2-(2- methoxyethoxy)ethanol (270 mL), and the mixture was stirred for about 0.5 hour before it was combined with a solution of 3a (12.0 g, 0.0640 mol) in 2-(2-methoxyethoxy)ethanol (335 mL). The stirred mixture was heated under N2 at 150-160°C for 7 hours. This mixture was allowed to cool to about 110°C while another solution of guanidine (one-half the previous amount) in 2-(2- methoxyethoxy)ethanol was prepared. The second guanidine solution was added, and heating at 150-160°C was resumed. After 5 hours, the mixture was allowed to cool, then evaporated in vacuo (<1 mm) to a viscous mixture. Addition of cold H2O (-500 mL) gave a crude solid, which was collected and dried in vacuo. The crude product mixture (11.4 g) was dissolved in DMF, and die solution was swirled witii silica gel (about 40 g of 60-200 mesh). Evaporation in vacuo as before gave a solid dispersion of crude product mixture and silica gel. The dispersion was pulverized, dried further in vacuo, then applied to a column (9 ¥ 50-cm) of silica gel (60- 200 mesh poured from CHCI3). Gravity elution by CHCl3-MeOH (95:5) followed, and fractions homogeneous in 4a (Rf -0.55 on TLC using CHC-3-MeOH, 5:1) were combined for evaporation to give pure 4a (3.3 g, 23% yield). Spectral data: mass, m/z 229, MH+ for C11H12N6; IH NMR (Me 2 SO-d6) d0.92 (t, 3, CH3), 1.62 (m, 2 CH2CH2CH 3 ), 3.20 (m, 2, CH 2 CH2CH3), 6.8-7.0 (br, 2, NH 2 ), 7.32 (br s, 2, NH 2 ), 8.78 (s, 1, C 7 -H). 2,4-^Diammo-5-propylpyrido[2,3-d]pyrimidine-6-carboxaldehyde. Raney Ni (1.2 g damp) was added with the aid of 95-97% HCO2H (7 mL) to a solution of 2,4-&amino-5- propylpyrido[2,3-d]pyrimidine-6-carbonitrile (500 mg, 2.19 mmol) in 95-97% HCO2H (5 mL). The stirred mixture was heated at 75-80°C for 1.5 h. Raney Ni was removed by filtration, and the filtrate was evaporated. The residue was dissolved in hot H2O (20 mL), and the solution was filtered, then cooled, and treated with concentrated NH 4 OH to pH 7 to cause solid to precipitate. The mixture was kept at 0-5°C overnight before the solid was collected and washed with cold H2O. This crude material was applied to a silica gel column and eluted with CHCl3-MeOH (7:1). Evaporation of the product fractions afforded the aldehyde; yield 8% (41 mg). Anal. Calcd. for CιιHι 3 N5θ-0.2H2θ: C, 56.26; H, 5.75; N, 29.82. Found: C, 56.26; H, 5.70, N, 29.72. MS, m/e 232, MH+; UV, lmax (e ¥ 10-3) in 0.1N HCl, 235 (22.9), 257 (19.5), 316 (9.15); pH 7 buffer, 234 (15.7), 265 (17.4), 318 (11.0), 346 (12.2); in 0.1N NaOH, 234 (17.5), 265 (16.6), 347 (12.8); IH NMR (Me 2 SO-d6), d 0.92 (t, 3, CH 3 ), 1.55 (sext, 2, CH2CH2CH3), 3.46 (t, 2, CH2CH2CH3), 6.84 (br s, 2, NH 2 ), 7.30 (s, 2, NH 2 ), 8.88 (s, 1, C7-H), 11.0 (s, 1, CHO). 2,4-Diamino-5-propylpyridol[2,3-d]pyriπ-idine-6-methanol. The aldehyde (95 mg, 0.41 mmol) was stirred with MeOH (20 mL), and the near-solution was treated with 3 portions of NaBF-4 (17 mg total, 0.45 mmol) added at 15-min intervals. Complete solution occurred after the first addition of NaBH4. The solution was left at 20-25°C for 1 h. The solution was treated with H2O (1 mL), neutralized (to pH 7) with glacial AcOH, and evaporated to near dryness. Solid residue was stirred with a little cold H2O (-1 mL), collected, and dried to give a first crop of 6-hydroxymethyl compound; yield 31% (30 mg); MS, m/e 234, MH+ for C11H15N5O: IH NMR (Me 2 SO-d6), d 0.96 (t, 3, CH3), 1.56 (sext, 2, CH2CH2CH3), 3.06 (t, 2, CH2CH2CH3), 4.52 (s, 2, CH 2 OH), 6.22 (s, 2, NH 2 ). 6.90 (s, 2, NH 2 ), 8.48 (s, 1, C7-H). The filtrate from this sample was evaporated to dryness, and the residue was extracted several times with boiling EtOAc to provide another crop of hydroxymethyl compound (80 mg), MS m/e 234. 6-(Bromomethyl)-2,4-diamino-5-propylpyrido[2,3-d]pyrimidine (1-6). The hydroxymethyl compound was treated with dry HBr in AcOH as reported for the methyl homolog (1-4). Addition of Et2θ caused precipitation of yellow sohd which was collected with the aid of Et2θ and dried in vacuo to afford the HBr salt of the product; MS m/e 296 and 298, MH+ for CnHi4BrN5, but with higher mass peaks present. The IH NMR spectrum of the product mixture showed expected singlets due to the CH2Br (4.92) and the C7-H (8.78). Relative integral values suggested about 10 mole-percent of the product. N-[5-[[(2,4-Diamino-5-propylpyrido[2,3-d]pyrimidin-6- yl)methyl]amino]thiophene-2-carbonyl]L-glutamic Acid (1-11). A mixture of the crude bromomethyl preparation described above (0.55 g), diethyl N-(5-aminothiophene-2-carbonyl)-L- glutamate (0.40 g, 1.2 mmol) and CaCO3 (160 mg) in Me2NAc (7 mL) was stirred at 20-25°C for 4 days. Insoluble material was removed by filtration, and the filtrate was evaporated to dryness. The residue was chromatographed on silica get with elution by CHCl3-MeOH (5:1) to give fractions homogeneous in the desired product. Evaporation of the pooled fractions gave the diethyl ester as a pale-orange solid (20 mg); MS m/e 545, MH+ for C25H33N7O5S. For ester hydrolysis, this sample was dissolved in 1 N NaOH (0.37 mL), and the solution was kept at 20- 23°C for 5 h. The solution was clarified (Celite mat), then acidified to pH 3.8-4.0 with 2 N HCl. After the mixture had been refrigerated, the solid was collected and dried; yield 6 mg. MS, m/e 488, MH+ for C21H25N7O5S. Example F*: Synthesis Of Compound 71, Table IIB, By Procedure III 2-Carboxythiophene-5-acetic Acid Dimethyl Ester (H-2). Freshly distilled dusopropylaπώie (24.6 g, 0.24 mol) in 250 mL of dry tetrahydrofuran (THF) was cooled to 0°C under argon then treated dropwise with 98 mL (0.24 mol) of 2.5M BuLi in hexane. After 1 h, the LDA solution was added dropwise with stirring to a -30° mixture of 15.0 g (0.11 mol) of 5- methylthiophene-2-carboxylic acid (II- 1) in 300 mL of dry THF. The temperature of the resulting red solution was allowed to rise to 0°C and was so maintained for another 2 h. Carbon dioxide was bubbled through the solution to produce a yellow precipitate. The mixture was stirred at ambient temperature for 2 h and filtered. The yellow filter cake was suspended in 300 mL of MeOH, and the mixture was cooled to 0°C and treated witii 100 mL of MeOH saturated with dry HCl. The mixture was stirred at room temperature for 72 h, concentrated in vacuo and the residue was partitioned between Et2θ (500 mL) and 250 mL of sat'd NaHCO3. TheEt2θ extract was washed with H2O (3 ¥ 250 mL), dried over MgS * and evaporated to leave a dark oil (15 g). Chromatography on flash silica gel (EtOAc-hexane, 1:19) gave 11.4 g of the product (51%) as a white, waxy solid; NMR (CDCI3) d 7.61 (d, IH, 3-H); 6.90 (d, IH,. 4-H); 3.87 (m, 5H, ArCOOCH3 + CH2); 3.82 (s, 3H, CH 2 COOCH 3 ); Anal. (C 9 H 10 O 4 S); Calc: C, 50.5; H, 4.71; Found: C, 50.6; H, 4.79. 5-[l-C^bometiιoxy-2-(2,4-diam o[2,3-d]pyrimidin-6-yl)ethyl]thiophene-2- carboxylic Acid Methyl Ester (H-3). The sodio derivative of 2-carbomethoxythiophene-5-acetic acid methyl ester 01- ) (12.0 mmol) was preparaed in DMF using NaH (480 mg of 60% dispersion in oil, 12.0 mmol). The mixture was treated with a solution of 1-3 (1.6 g, 4.0 mmol) in 10 mL of DMF at -25°C. The mixture was kept at - 10°C for one hour and then stirred for one hour at room temperature, followed by neutralization with solid CO 2 . Silica gel was added followed by evaporation in vacuo. The solid residue was applied to a silica gel column and the product eluted with CHCl3-MeOH (9:1) to afford II-3 in 23% yield (350 mg); mass spectrum, m/e 388, MH+ for C17H17N5O4S. 5-[l-Carboxy-2-(2,4-diammo[2,3-d]pyrimidin-6-yl)ethyl]thiophene-2-carboxyhc Acid (π-4). The ester II-3 (350 mg) was hydrolyzed by 4N NaOH (0.5 mL) in dimethylsulfoxide (DMSO) for 20 hours followed by removal of DMSO at 40° in vacuo. Precipitation of II-4 from a solution of its Na salt in H2O by treatment with IN HCl to pH 4.0 gave a gel-like precipitate. The gel became particulate solid after the mixture had been frozen solid and then allowed to thaw. The solid was then easily collected by filtration; yield 85% (275 mg); mass spectrum, m/e 360, MH+ for C 1 5H13N5O4S. Assay by HPLC, >93% purity. 5-[2-(2,4-Diamino[2,3-d]pyrimidin-6-yl)ethyl]thiophene-2-carboxylic Acid (II- • 5). A solution of II-4 (235 mg, 0.65 mmol) in DMSO (8 mL) was kept at 120-125°C for 20 min. Removal of DMSO in vacuo (<1 mm, bath to 40°C) gave E-5; mass spectrum, m/e 316, MH+ for C14H13N5O2S. This material was used directly for conversion to II-6. N-[5-[2-(2,4-Diamino[2,3-d]pyrimidin-6-yl)ethyl]thiophene-2-carbonyl]-L- glutamic Acid Diethyl Ester (II-6). A stirred mixture of II-5 (210 mg, 0.66 mmol) in DMF (25 mL) was treated with E13N (0.37 mL, 26 mg, 2.64 mmol) followed by i-BuOCOCl (0.09 mL, 92 mg, 0.66 mmol), then stirred at 20-23°C for 15 min before diethyl L-glutamate » HCl (158 mg, 0.66 mmol) was added. Three more additions of i-BuOCOCl (0.33, 0.17, and 0.17 mmol) were made at intervals of 15 min, and each was followed 1 min later by an equimolar amount of diethyl L-glutamate»HCl. The course of the conversion was followed by TLC (CHCl3-MeOH, 3 : 1), and a chromatogram observed 2 h after the final addition revealed one major UV-absorbing spot: DMF was then removed (<1 mm, 25-30°C). The residue was dissolved in MeOH, and the solution was treated with silica gel (2.0 g of 60-200 mesh). Evaporation gave a dry dispersion of crude product in silica gel which was applied atop a column of silica gel. The column was eluted with CHCl3-MeOH (5 : 1), and fractions were examined by TLC. Appropriate fractions were combined and evaporated, and the residue was stirred with Et2θ, then collected by filtration. The yield of diethyl ester (E-6) was 36% (120 mg); mass spectrum, m/e 501, MH+ for C23H28N6O5S. N-(5-(2-(2,4-Diamino[2,3-d]pyrimin-6-yl)thiophene-2-carbonyl]-L-glutamic Acid (π-7). The ester (II-6, 120 mg) was dissolved in MeOH (10 mL) along with IN NaOH (0.5 mL). After 2 days at 20-23°C in a stoppered flask protected from light, the solution was evaporated in vacuo, bath 25°C, to remove MeOH which was replaced with H2O (10 mL). More IN NaOH (0.25 mL) was added, and the aqueous solution was left at 20-23°C for 2 days before it was treated with IN HCl (pH 4.0). The precipitated product was collected, washed with H2O, and dried (in vacuo) to afford E-7 in 37% yield (43 mg); MS, m/e 445, MH+. Anal. Calcd for Ci9H2θN6θ5S » 2H 2 O: C, 47.49; H, 5.03; N, 17.49. Found: C, 47.23; H, 4.81; N, 17.13. Example G': Synthesis Of Compound 93, Table π, By Procedure IV 3-Carboxypyridine-6-acetic Acid Dimethyl Ester (III-2). This diester was similarly prepared as for II-2 from 6-methylnicotinic acid (III-l) as a yellow solid, mp 56-57°C; NMR (CDCI3) d 9.10 (m, IH, 6-H); 8.21 (m, IH, 4-H); 7.33 (m, IH, 3-H); 3.84 (m, 8H, CH 2 COOCH3dArCOOCH 3 ); Anal. (CιoHnNO 4 ); Calc: C, 57.4; H, 5.30; N, 6.70. Found: C, 57.5; H, 5.33; N, 6.54. 10-Carbomethoxy-4-deoxy-4-amino-5,10-dideaza-3'-azapteroic Acid Methyl Ester CM-3). Alkylation of the sodio derivative of 3-carbomethoxy-6-pyridylacetic acid methyl ester (HI-2) with 1-3 to produce III-3 was done as described for the preparation of II-3. The product obtained in greater than 90% yield was nearly homogeneous by TLC (CHC-3-MeOH, 5: 1) and produced the expected mass spectral peak of m/e 393, MH+ for C18H18N6O4. Purity assay by HPLC showed the main component to be >86% with respect to UV-absorbing material. This material was used as such for conversion to IH-4. 10-Carboxy-4-deoxy-4-amino-5,10-dideaza-3'-azapteroic Acid (III-4). Ester hydrolysis of IE-3 was conducted as described for the conversion of II-3 to II-4. The overall yield of II-4 from 1-3 was 86% (1.25 g from 1.66 g, 4.08 mmol of 1-3). Mass spectrum, m/e 355, MH+ for Ci6Hi4N 6 O . 4-Deoxy-4-ami-no-5,10-dideaza-3'-azapteroic Acid (III-5). Compound III-4 (1.25 g) was suspended in DMF (30 mL), and the stirred mixture was heated at 75°C for 20 min. Evolution of CO2 appeared to cease after 10 min although solution did not occur. DMF was removed in vacuo, and the residue was stirred with H2O, collected and dried to give IH-5 (0.88 g, -84% yield); mass spectrum, m/e 311, MH+ for C15H 14 N6O2. N-[2-[2-[(2,4-Diaminopyrido[2,3-d]pyrimidin-6-yl)ethyl]-5-pyridyl]carbonyl3-L- glutamic Acid (J L-7). (5,10-Dideaza-3'-azaaminopterin). Compound IH-5 was coupled with diethyl L-glutamate in the same manner as described for compound II-6 (Example F). The yield of pure ester, homogeneous by TLC (CHC- 3 -MeOH, 5 : 1; Rf -0.5), was about 18% (215 mg from 740 mg, 2.38 mmol of IH-5); Mass spectrum, m/e 496, MH+ for C24H29N7O5. Hydrolysis of the ester groups of this product (175 mg, 0.353 mmol) as described under the preparation of II-7 led to pure IH-7«3H2θ in 77% yield (119 mg), Mass spectrum, m/e 440; IH NMR d 1.95, 2.08 (two m, 2, CHCH2 nonequivalent), 2.34 (t, 2, CH2CO), 3.06 (m, 2, C9H2 or C10H2), 3.17 (m, 2, C9H2 or CιoH 2 ), 4.38 (m, 1, CHCH 2 ), 6.85 (s, 2, NH 2) , 7.36 (d, 1, pyridyl- 3-H), 7.84 (s, 2, NH 2 ), 8.12 (m, 1, pyridyl-4-H), 8.37 (d, 1, C5-H), 8.54 (d, 1, C7-H), 8.68 (d, 1, CONH), 8.97 (d, 1, pyridyl-6-H between N and carboxamide). Anal. Calcd for C2θH2i 7θ 5 ^3H 2 O: C, 48.68; H, 5.51; N, 19.87. Found: C, 48.45; H, 5.55; N, 19.75. Example H 1 : Synthesis Of Compound 99, Table HB, By Procedure IV 10-Carbomethoxy-4-deoxy-4-amino-5-methyl-5, 10-dideaza-3'-azapteroic Acid Methyl Ester (DI-8). NaH (480 mg of 60% in oil, 12.0 mmol) was suspended in DMF (12 mL), and the mixture was chilled to 0°C, then treated with a solution of 3-carbomethoxy-6- pyridylacetic acid methyl ester (m-2, 2.50 g, 12.0 mmol) in DMF (12 mL). After 0.5 h at 0°C, the stirred mixture was chilled to -25°C, treated with a solution of I-4«I.7HBr»0.5AcOH (1.71 g, 3.92 mmol) in DMF (12 mL), then allowed to warm to -10°C. After 1 h at near -10°C, the solution was allowed to warm to ambient temperature. After 1 h at 20-23°C, the mixture was neutralized by the addition of small pieces of solid CO 2 . Addition of silica gel (7.5 g of 60-200 mesh) followed, and the resulting mixture was evaporated to dryness (<1 mm, bath to 40°C) to give a dispersion of crude product in silica gel which was applied to a column of silica gel. Elution by CHC-3-MeOH (9 : 1) led to fractions homogeneous in III-8 according to TLC (CHC-3-MeOH, 3: 1; Rf -0.6). Evaporation of the combined fractions gave III-8 in 43% yield (668 mg); mass spectrum, m/e 397, MH+ for C19H20N6O 4 . 10-Carboxy-4-deoxy-4-amino-5-methyl-5, 10-dideaza-3'-azapteroic Acid (IH-9). A suspension of πi-8 (668 mg, 1.69 mmol) in DMSO was treated with 4N NaOH (1.0 mL). The resulting clear solution was kept under N2 in a stoppered flask protected from light for 20 h. After the solvent had been removed by short-path distillation in vacuo (<1 mm, bath to 40°C) the residue was dissolved in H2O (30 mL), and the filtered solution was acidified to pH 5 using glacial AcOH. The mixture was kept several hours in a refrigerator before the solid was collected, washed with H2O, and dried in vacuo (over P2O5). This material was found through HPLC assay results and mass spectral data to be a mixture consisting of 88% compound πi-9 (m/e 369, MH+ for C 1 7H16N6O4) and 12% of the sequential product HMO (m/e 325 r MH+ for C16H16N6O2). The weight of the mixture (594 mg) corresponds to conversion of 97%. 4-Amino-4-deoxy-5-methyl-5,10-dideaza-3'-azapteroic Acid (HI- 10). The above material πi-9 (594 mg) was suspended in DMF (15 mL), and the stirred mixture was kept at 60- 65°C for 12 min. Decarboxylation was slow at this temperature, and the bath was raised to 80- 85°C. Heating was continued 15 min longer. To ensure complete reaction, DMF was removed in vacuo and replaced with DMSO. The resulting clear solution was kept at 60-65°C for 15 min, then examined by HPLC showing complete decarboxylation. DMSO was removed in vacuo, and the product was precipitated from a clarified (Norit, Celite) aqueous solution of its Na salt by addition of AcOH to pH 5; yield 77% (443 mg); Mass spectrum, m/e 325, MH+. Anal. Calcd for C16H16N6O2: C, 54.69; H, 5.45; N, 23.92. Found: C, 54.77; H, 5.13; N, 24.41. N-[2-[2-[(2,4-Diamino-5-methylpyrido[2,3-d]pyrimidin-6-yl)ethyl]-5- pyridyl]carbonyl]-L-glutamic Acid Diethyl Ester (III-ll). A stirred mixture of III-10 (382 mg, 1.09 mmol) in DMF (40 mL) was treated with Et3N (0.61 mL, 0.44 mg, 44 mmol) followed by i- BuOCOCl (0.14 mL, 0.15 g, 1.08 mmol), then stirred at 20-23°C for 15 min before diethyl L- glutamate » HCl (261 mg, 1.09 mmol) was added. Three more additions of i-BuOCOCl (0.55, 0.27, and 0.27 mmol) were made at intervals of 15 min, and each was followed 1 min later by an equimolar amount of diethyl L-glutamate»HCl. The course of the conversion was followed by TLC (CHCl3-MeOH, 3:1), and a chromatogram observed 2 h after the final addition revealed one major UV-absorbing spot. DMF was then removed (<1 mm, 25-30°C). The residue was dissolved in MeOH, and the solution was treated with silica gel (2.0 g of 60-200 mesh). Evaporation gave a dry dispersion of -crude product in silica gel which was applied atop a column of silica gel (-300 mL of 230-400 mesh). The column was eluted with CHCl3-MeOH (5:1), and fractions were examined by TLC. Appropriate fractions were combined and evaporated, and the residue was stirred with Et2θ, then collected by filtration. The air-dried sample was stirred with H2O, and the H2O-insoluble solid was dried to give the diethyl ester (IE-11) (281 mg, 47% as dihydrate). Mass spectrum, m/e 510, MH+. Anal. Calcd for C25H 3 ιN7θ5«2H 2 O: C, 55.04; H, 6.47; N, 17.97. Found: C, 55.01; H, 6.23; N, 17.88. N-[2-[2-[(2,4-Diamino-5-methylpvrido[2,3-d]pyrimidin-6-yl)ethyl]-5- pyridyl]carbonyl[-L-glutamic Acid (HI-12). (5-Methyl-5,10-dideaza-3'-azaaminopterin). The ester (261 mg, 0.478 mmol) was dissolved in MeOH (20 mL) containing IN NaOH (1.1 mL). After 2 days at 20-23°C in a stoppered flask protected from light, the solution was evaporated in vacuo, bath 25°C to remove MeOH which was replaced with H2O (20 mL). More IN NaOH (0.5 mL) was added, and the aqueous solution was left at 20-23°C for 2 days longer before it was treated with IN HCl to pH 4.0 to cause precipitation. The product was collected, washed with H2O, and dried to give HI-12 in 88% yield (204 mg), Mass spectrum, m/e 454, MH+; UN (lmax, e¥ 10-3) 0.1ΝHC1, 228 (38.6), 270 (17.6), 319 (7.96); pH 7, 233 (38.5), 268 (16.6), 334 (6.01); 0.1Ν NaOH, 233 (37.4), 269 (18.1), 345 (6.72); 1HNMR (Me 2 SO-d6) d 1.95, 2.06 (two m, 2, CHCH2 nonequivalent), 2.34 (t, 2, CH 2 CO), 2.68 (s, 3, CH3), 3.06 (m, 4, C9H2C 1 0H2), 4.38 (q, 1, CHCH 2 ), 6.90 (s, 2, NH 2 ), 7.34 (m, 3, NH 2 overlapping pyridyl-3-H), 8.10 (m, 1, pyridyl-4-H), 8.30 (s, 1, C7-H), 8.64 (d, 1, CONH), 8.96 (d, 1, pyridyl-6-H; between pyridyl N and carboxamide). Anal. Calcd for C2iH28N7θ5»1.6H2θ: C, 52.30; H, 5.48; N, 20.33. Found: C, 52.19; H, 5.34; N, 20.45. The deazaaminopterin compound can be administered per se, or in association with a pharmaceutically acceptable diluent or carrier. The invention accordingly also provides a pharmaceutical composition in dosage unit form comprising from 0.1 to about 500 mg of the deazaaminopterin compound, per dosage unit, together with a pharmaceutically acceptable nontoxic inert carrier or diluent therefore. The deazaaminopterin compound can be used as such, or in the form of an acid addition salt. These salts are formed with one or more free NH2 groups of the deazaaminopterin molecule. Typically, the compounds are injected in the form of their sodium salts in aqueous solution. Other salts, e.g., K, Ca, NH4, etc. could be used as prepared from the appropriate hydroxide or carbonates. The acid addition salts are preferably the pharmaceutically acceptable, nontoxic addition salts with suitable acids, such as those with inorganic acids, for example, hydrochloric, hydrobromic, nitric, sulphuric, and phosphoric acids; and with organic acids, such as organic carboxylic acids, for example, glycolic, maleic, hydroxymaleic, malic, tartaric, citric, calicylic, o-acetyloxybenzoic, nicotinic, and isonicotinic acid; and organic sulphonic acids, for example, methanesulphonic, ethanesulphonic, 2-hydroxyethanesulphonic, toluene-p-sulphonic, and naphthalene-2-sulphonic acid. An acid addition salt can be converted into the free compound according to known methods, for example, by treating it with a base, such as with a metal hydroxide or alkoxide, for example, an alkali metal or alkaline earth metal hydroxide, for example, lithium hydroxide, sodium hydroxide, potassium hydroxide or calcium hydroxide; with a metal carbonate, such as an alkali metal or an alkaline earth metal carbonate or hydrogen carbonate, for example, sodium, potassium or calcium carbonate or hydrogen carbonate, with ammonia; or with a hydroxyl ion exchange resin, or with any other suitable reagent. An acid addition salt may also be converted into another acid addition salt according to known methods; for example, a salt with an inorganic acid may be treated with a metal salt, for example a sodium, barium or silver salt, of an acid in a suitable diluent, in which a resulting inorganic salt is insoluble and is thus removed from the reaction medium. An acid-addition salt may also be converted into another acid addition salt by treatment with an anion exchange preparation. The glutamic acid COOH groups can also be in salt form, as the ammonium NH 4 , alkali metal salts (Na"*" 1 ", K + ), or the nontoxic alkaline earth metal salts (Ca ++ ) of the glutamate COOH groups. The deazaaminopterin compound or salt thereof can be administered to the animal by any available route, including oral and parenteral (intravenous, intraperitoneal, subcutaneous, and intramuscular) administration. The amount administered is sufficient to ameliorate the arthritis or other proliferative disease, and will depend upon the type of arthritis, the species of animal, and the weight of the animal. For example, in human administration, a dosage of deazaaminopterin compound within the range from about 0.1 mg/kg to about 500 mg kg per day should be sufficient. Dosages in the higher part of the range, approaching 500 mg/kg, are normally administered in conjunction with leucovorin (dl-r-formyl tetrahydrofolate) to reduce toxicity. In the treatment of lower test animals, a similar dosage range is therapeutic. The upper limit of dosage is that imposed by toxic side effects, and can be determined by trial and error for the animal to be treated, including humans. To facilitate administration, the deazaaminopterin compound or salt thereof can be provided in composition form, and preferably in dosage unit form. While the compound can be administered per se, it is normally administered in conjunction with a pharmaceutically acceptable carrier therefor, which dilutes the compound and facilitates handling. The term "pharmaceutically acceptable" means that the carrier (as well as the resulting composition) is sterile and nontoxic. The carrier or diluent can be solid, semisolid, or liquid, and can serve as a vehicle, • excipient, or medium for the compound. Exemplary diluents and carriers include lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, mineral oil, cocoa butter, oil of theobroma, alginates, tragacanth, gelatin, syrup, methyl cellulose, polyoxyethylene sorbitan monolaurate, methyl- and propylhydroxybenzoate, talc, or magnesium stearate. For convenience in handling, the deazaaminopterin compound and carrier or diluent can be enclosed or encapsulated in a capsule, sachet, cachet, gelatin, paper, or other container, especially when intended for use in doage units. The dosage units can for example take the form of tablets, capsules, suppositories, or cachets. The following Examples 1-7 illustrate various forms of dosage units in which the deazaaminopterin compounds or salts thereof can be prepared: Example 1 Tablet Formation Mg/tablet Deazaaminopterin compound 15 Lactose 86 Corn starch (dried) 45.5 Gelatin 2.5 Magnesium stearate 1.0 The deazaaminopterin compound is powdered and passed through a mesh sieve and well mixed with the lactose and 30 mg of the corn starch, both passed through a sieve. The mixed powders are massed with a warm gelatin solution, prepared by stirring the gelatin in water and heating to form a 10% w/w solution. The mass is granulated by passing through a sieve, and the moist granules dried at 40°C. The dried granules are regranulated by passing through a seive and the balance of the starch and the magnesium stearate is added and tiioroughly mixed. The granules are compressed to produce tablets each weighing 150 mg. Examnle 2 Tablet Formation Mg/tablet Deazaaminopterin compound 100 Lactose 39 Corn starch (dried) 80 Gelatin 4.0 Magnesium stearate 2.0 The method of preparation is identical with that of Example 1 , except that 60 mg of starch is used in the granulation process and 20 mg during tableting. Exam l 3 Capsule formation Mg/capsule Deazaaminopterin compound 250 Lactose 150 The deazaaminopterin compound and lactose are passed through a sieve and the powders well mixed together before filling into hard gelatin capsules of suitable size, so that each capsule contains 400 mg of mixed powders. Example 4 Suppositories Mg/suppositories Deazaaminopterin compound 50 Oil of Theobroma 950 The deazaaminopterin compound is powdered and passed through a sieve and triturated with molten oil of theobroma at 45°C to form a smooth suspension. The mixture is well stirred and poured into molds-, each of nominal 1 g capacity, to product suppositories. Examples Cachets Mg cachet Deazaaminopterin compound 100 Lactose 400 The deazaaminopterin compound is passed through a mesh sieve, mixed with lactose previously sieved and fitted into cachets of suitable size so that each contains 500 mg. Example 6 Intramuscular injection (sterile suspension in aqueous vehicle) Mg Deazaaminopterin compound 10 Sodium citrate 5.7 Sodium carboxymethylcellulose (low viscosity 2.0 grade) Methyl para-hydroxybenzoate 1.5 Propyl para-hydroxylbenzoate 0.2 Water for injection to 1.0 ml Example 7 Intraperitoneal intravenous or subcutaneous injection (sterile solution in aqueous carrier system) Mg Deazaaminopterin compound, hydrochloric acid addition salt 15 Sodium citrate 5.7 Sodium carboxymethylcellulose (low viscosity 2.0 grade) Methyl para-hydroxybenzoate 1.5 Propyl para-hydroxylbenzoate 0.2 Water for injection to 1.0 ml Example 8: Tn Vivo Biology Of Tvoe TI Collagen Arthritis And Methotrexate Treatment Using 5-Deazaaminopterin Compounds Tn Table IA Of Formula IT Where A Is The following data illustrate administration to mice of several deazaaminopterin compounds of the invention and methotrexate in the evaluation of anti-inflammatory activity. The data are presented as two separate observations, the visually observed presence of inflammation in the mouse, and the caliper-measured degree of swelling of the rear paws of the mouse. The efficacy evaluation used a mouse model of inflammatory disease that occurs in response to an antigenic challenge with Type E collagen [J. S. Courtenay et al., Nature.283. 666-668 (1980)]. The fundamental aspects of the model allow it to serve as a representative presentation of human disease. The parallels between the known aspects of the mouse model and rheumatoid arthritis include a humoral response in which antibodies are produced to an antigen that is present in the joint tissue and the antigenic challenge is accompanied by cell-mediated aspects of immunity. The resultant inflammation of the joint tissue yields facets of periostitis, synovial lining hyperplasia, degradation of bone and cartilage and pannus and new bone formation. The basic elements of the model included the immunization of DBA/1 mice with a suspension of fetal bovine Type II collagen (1 mg ml) prepared in complete Freund's adjuvant. The primary injection was given using 0.1 ml of the collagen emulsion giving a total of 0.1 mg of Type II collagen per mouse. The animals were then given a booster injection of Type E collagen (100 μg in 0.01 M acetic acid ) on day 21 by intraperitoneal injection. The results of the in vivo testing of methotrexate showed that using prophylactic regimens in which drug was begun two days prior to administration of antigen (Type E collagen) was more effective than starting drug at day 19, two days prior to the first and only boost with Type E collagen. Typically, in this model the untreated positive control animals have an incidence of arthritis ranging from 90 to 100% of injected animals at day 44. The effect of methotrexate and test compounds on the extent of inflammation was determined by direct analysis of paw swelling using caliper measurements. The results are presented in Table EA, and show a direct correlation between the decrease in the number of animals having disease and a decrease in the extent of inflammation, as determined by paw swelling. TaMe ΠA No Mice on day Ave. thickness of indicated 0 rear paws (mm) over dav S 30-44g Dose Compound fmg/kiJ) Dav 30 Day 37 Dav 44 Treated Untreated None 31/43 38/43 41/43 5-Me-10-H 1.5 0.8 2/8 6/8 (2.18-2.55) (2.33-2.87) (5-Me-5-DA) 5-Me-10-Me 1.5 1/8 4/8 5/8 (2.26-2.44) (2.33-2.87) (5-Me-5-DMTX) 5-EMO-H 1.0 2/8 5/8 2/8 (2.19-2.24) (2.56-2.98) (5-Et-5-DA) 5-Et-10-Me 0.75 0/7 1/7 1/7 (2.20-2.18) (2.24-2.63) (5-Et-5-DMTX) 5-Pr-10-H 1.5 0/7 0/7 0/7 (2.14-2.15) (2.24-2.63) (5-Pr-5-DA) 5-H-10-Propargyl 12.0 3/8 2/8 6/8 (2.22-2.37) (2.34-2.78) (10-Prgl-5-DA) 5-Me-10-Propargyl 1.5 3/8 3/8 2/8 (2.29-2.29) 2.56-2.98) (5-Me-10-Prgl-5-DA) 5-Me-lO-Allyl 3.0 0/8 0/8 0/8 (2.12-2.16) (2.32-2.72) (5-Me-10-Allyl-5-DA) MTXa 9.0 1/22 1/22 6/22 (2.18-2.34) (2.24-2.98) a Methotrexate (MTX) and untreated controls are composites from multiple runs. * » Visual evidence of inflammation. c Values in parentheses are 30 day and 44 day measurements vs. equivalent for untreated controls; decrease in inflammation vs. control is most notable at day 44. It is apparent from the above results that the number of test mice affected was very considerably descreased by administration of deazaaminopterin compound. The results show that deazaaminopterin compound on a similar dosage level to be at least as effective as methotrexate, and since methotrexate is accepted as effective the deazaaminopterin compound is to be expected to be at least as effective as methotrexte, under similar conditions. The potent anti-arthritic activity of the deazaaminopterin compounds tested is evident from the results. In addition to an effectiveness at least as great methotrexate at a similar dosage level, the compounds of the invention have an advantage in a lower-toxicity, meaning that dosages higher than methotrexate can be possible. The data in Table EIA show less, cytotoxicity on a human Uver cell Une (Chang liver) than methotrexate, in terms of the rates of cytotoxic potency versus methotrexate, whose ratio in those terms is 1.00. Thus, the higher the ratio is above 1, the lower the toxicity of the compound in respect to methotrexate. Table IIIA Ratio Of Cvtotoxic Potency Versus Methotrexate For Inhibition Of Human Liver Cell Growth In Culture Compound Ratio MTX 1.00 5-Me-10-NH 4.39 5-Me-lO-NMe 2.79 5-Et-lO-NH 4.50 Taftlg lllA Ratio Of Cvtotoxic Potency Versus Methotrexate For Inhibition Of Hunrøη Liver Cell Gro h In CyUwr Compound Ratio 5-EMO-NMe 3.18 5-Pr-10-NH 1.78 5-Bu-lO-ΝH 7.20 5-Me-lO-N-Propargyl 21.18 5-H-10-N-Propargyl 1.26 5-Me-10-N-allyl 5,10-Dideazaaminopterin 2.46 10-Me-5,10-Didea2aaminopterin 2.69 Example 9: In Vivo Biology Of Type II Collagen Arthritix And Methotrexate Treatment Using 5-Deazaaminonferin Compounds In Table IA Of Formula IT Wherein A JsJϋ The following illustrates administration to mice of several 5-deazaaminopterin compounds of the invention and methotrexate in the evaluation of anti-inflammatory activity. The data are presented in Table IVA below as two separate observations, the visuaUy observed presence of inflammation in the mouse, and the visuaUy observed presence of swelling of the rear paws of the mouse. The efficacy evaluation uses a mouse model of inflammatory disease that occurs in response to an antigenic chaUenge with Type E collagen [J. S. Courtenay et al., Nature.283. 666-668 (1980)], as described in Example 8 hereinabove. Table IVA Visual Observation Of Inflammation And Paw Swelling Visual Observation of Presence Compound Dose (mg/kg) Inflammation of Swelling None 5-Me-10-H 1.5 Yes Yes 5,10-dideaza 5-Me-lO-Me 1.5 Yes Yes 5,10-dideaza 5-Et-lO-H 1.0 Yes Yes 5,10-dideaza 5-Et-10-Me 0.75 Yes Yes 5,10-dideaza 5-Pr-10-Me 1.5 Yes Yes 5,10-dideaza 5-H-10- 12.0 Yes Yes Propargyl 5,10-dideaza 5-Me-10- 1.5 Yes Yes Propargyl 5,10-dideaza 5-Me-lO-Allyl 3.0 Yes Yes 5,10-dideaza The results show the deazaaminopterin compound on a similar dosage level to be at least as effective as methotrexate, and since methotrexate is accepted as effective the dea∑aaminopterin compound is to be expected to be at least as effective as methotrexate, under similar conditions. The anti-arthritic activity of the deazaaminopterin compounds tested was confirmed by the results. Example 10: In Vivo Biology Of Type TT Collagen Arthritis And Methotrexate Treatment Using Heteroarovl-5-DeazaaminonterTn Compounds Nos. A' To H' Of Table TB The foUowing data illustrate administration to mice of Compounds Nos. A' to H 1 of Table IB of the invention and methotrexate in the evaluation of anti-inflammatory activity. The data are presented as two separate observations, the visually observed presence of inflammation in the mouse, and the caliper-measured degree of sweUing of the rear paws of the mouse. The efficacy evaluation used a mouse model of inflammatory disease that occurs in response to an antigenic challenge with Type E collagen [Courtenay et al., Nature. 283. 666- 668 (1980)], as described in Example 8 hereinabove. The effect of methotrexate and test compounds on the extent of inflammation was determined by direct analysis of paw swelling using caliper measurements. The results are presented in Table EB, and show a direct correlation between the decrease in the number of animals having disease and a decrease in the extent of inflammation, as determined by paw swelling. Table IIB No mice affected on day indicated 0 Avg. thickness of rear paws (mm) over days 30-44C Compound Dose (mg kg) Day 30 Day 37 Day 44 Treated Untreated None 31/43 38/43 41/43 2.29-2.73 A' 6.0 0/5 2/5 2/5 2.19-2.33 6.0 0/8 1/8 3/8 2.13-2.27 6.0 2/8 4/8 7/8 2.13-2.32 8.0 2/8 7/8 7/8 2.19-2.63 12.0 5/8 7/8 7/8 2.28-2.67 9.0 1/22 1/22 6/22 2.128-2.34 a MTX and untreated controls are composites from multiple runs. b Visual evidence of inflammation. c Values in parentheses are 30 day and 44 day measurements vs. equivalent for untreated controls; decrease in inflammation vs. control is most notable at day 44. It is apparent from the above results that the number of test mice affected was very considerably decreased by administration of heteroaroyl-5-deazaaminopterin or 5,10- dideazaaminopterin compound. The results show that heteroaroyl-5-deazaaminopterin or 5,10- dideazaaminopterin compound on a similar dosage level to be at least as effective as methotrexate, and since methotrexate is accepted as effective the heteroaroyl-5- deazaaminopterin or 5,10-dideazaaminopterin compound is to be expected to be at least as effective as methotrexate, under similar conditions. The potent anti-arthritic activity of the heteroaroyl-5-deazaaminopterin or 5,10-dideazaaminopterin compounds tested is evident from the results.

Claims: WHATTSCI.ATMF.DTS. 1. 5-deazaaminopterin and 5,10-dideazaaminopterin compounds represented by the formula: CH 2 COOH wherein AisCHorN; X is one of and Ri is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms; and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms; provided that when X is O and A is N, then Ri is alkyl, alkenyl, or alkynyl and when Ri is alkyl, then R2 is hydrogen or alkenyl, and further provided that when X is O and A is CH and Ri is hydrogen or alkyl, then R2 is alkenyl or alkynl. 2. The compounds of Claim 1 wherein X is one of and Ri is hydrogen or alkyl having from one to about eight carbon atoms; and R 2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms. 3. The compounds of Claim 2 wherein Ri is hydrogen or alkyl having from one to three carbon atoms, and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to three carbon atoms. 4. The compounds of Claim 1 wherein X is one of and Ri is hydrogen or alkyl having from one to about eight carbon atoms; and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms. 5. The compounds of Claim 4 wherein Ri is hydrogen or alkyl having from one to about three carbon atoms, and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about three carbon atoms. The compounds of Claim 1 wherein X is 0 and A is CH, and at least one of Ri and R2 is independently alkyl, alkenyl, or alkynyl having from three to about five carbon atoms. The compounds of Claim 1 wherein X is O and A is N, and at least one of Ri and R2 is independently alkyl, alkenyl, or alkynyl having from three to about five carbon atoms. 8. The compounds of Claim 7 wherein X is and at least one of Ri and R2 is alkyl having from three to about five carbon atoms. 9. The compounds of Claim 8 wherein the alkyl is propyl or butyl. 10. The compounds of Claim 1 wherein X is and at least one of Ri and R 2 is alkenyl. 11. The compounds of Claim 10 wherein the alkenyl is allyl or butenyl. 12. 5-deazaaminopterin and 5, 10-dideazaaminopterin compounds represented by the formula: COOH wherein Ais CHorN; X is one of and Ri is alkyl, alkenyl, or alkynyl having from three to about eight carbon atoms; and R2 is alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms. 13. The compounds of Claim 12 wherein X is and A is N, and at least one of Ri and R 2 is independently alkyl or alkenyl having from three to about five carbon atoms. 14. The compounds of Claim 1 wherein X is 1 o and one of Ri and R2 is alkyl and the other of Ri and R2 is alkenyl. 15. The compounds of Claim 1 wherein X is o and one of Ri and R 2 is alkyl and the other of Ri and R 2 is alkynyl! 16 A method of treating arthritis and other proliferative diseases which comprises administering to a warm-blooded animal having an inflammation of the joints or other evidence of the diseases, a therapeutic and relatively nontoxic amount of a 5- deazaaminopterin or 5,10-dideazaaminopterin compound having the formula: CH 2 COOH wherein A is CH or N; X is one of andRi is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms; and R2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms. 17. The method of Claim 16 wherein the compound is administered as a pharmaceuticaUy acceptable salt thereof. 18. The method of Claim 16 or 17 wherein the compound is administered in an amount within the range from about 0.1 to about 500 mg per day. 19. The method of Claim 16 or 17 wherein the compound is administered with an inert diluent or carrier. 20. The method of Claim 16 or 17 the compound is administered orally or parenteraUy. 21. A pharmaceutical composition in dosage unit form for treating arthritis or other proliferative disease comprising an amount within the range from about 0.1 to about 500 mg per dosage unit therapeuticaly effective to ameliorate arthritis or other proliferative disease of a 5-deazaaminopterin or 5,10-dideazaaminopterin compound together with a pharmaceutically acceptable nontoxic carrier or diluent thereof; the compound having the formula: CH, COOH wherein A is CH or N; X is one of and Ri is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms; and R 2 is hydrogen or alkyl, alkenyl, or alkynyl having from one to about eight carbon atoms. 22. The pharmaceutical composition of Claim 13 which is in the form of a pharmaceutically acceptable acid addition salt. 23. The pharmaceutical composition of Claim 21 or 22 in sterile aqueous, aqueous dispersion, capsule, cachet, or suppository form.

Inventors: COLWELL WILLIAM T, DEGRAW JOSEPH I, PIPER JAMES R, SIROTNAK FRANCIS M, SMITH R LANE, COLWELL, WILLIAM, T., DEGRAW, JOSEPH, I., PIPER, JAMES, R., SIROTNAK, FRANCIS, M., SMITH, R., LANE
Applicants: STANFORD RES INST INT, SRI INTERNATIONAL
Application year: 1993
Main classification: C07D471
IPC codes: A61K31, A61P29, C07D471