JPS58185584A - Pyrido(2,3-d)pyrimidine - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to
Methylamine]benzoyl)-L-glutamic acid (5-
deazafolate), N-1a-[[(2-amino-4(3H
)-Oquinpyrito [2゜3-d]-pyrimidine-6-
yl)methyl]methylamine]ben/yl 1-L-glutamic acid (5-deaza-NIQ-methylfolate), N-(
4-[:2゜4-noaminobiri)"[2,3-d]pyriminon-6-yl)methylamine]benzoyl]-L-
Glutamic acid (5-deazaaminotutilin), N-(4
~[C(2,4-diaminopyridoC2,3-d]biriminon-6-yl)methyl]methylamine]penozoyl)
-L-glutamic acid (5-7'7 demethotrexate)
The present invention also relates to intermediates useful in the preparation of pyrido'(2,3-d]pyriminone containing pyrido'(2,3-d)pyriminone. It also relates to l 0-substituted folic acids and methods for producing such novel compounds.

Potent sohydrofolate reductase inhibitors, such as aminotutilin and methotrexate, are known antifolate agents useful in the prevention and treatment of acute leukemia and related conditions. These aminotutilins and methotrexate have as their main mechanism of action competitive inhibition of the enzyme dihydrofolate reductase. In the process of DNA synthesis and cell regeneration, folic acid and its 7,8-dihydro derivatives must be reduced to tetrahuman sulfate by this enzyme. Antifolate activity such as aminopterin and methotrexate
Compounds with Ctivity ) suppress the reduction of both folic acid and 7°8-dihydrofolate and interfere with tissue cell regeneration.

Several types of quinazolinyl (5,8-dideadepterinonyl) analogs of folic acid, aminotutilin, and methotrexate are active in dihydrofolate reductase and thymidylate synthetase (thymidylate 5ynt).
hetase) (A, H, Calvert, T, R.

Noyones, P. J., Dardy, B., Grzelakowska, J. HMHK.
nes, P., J., Dady.

B., Grzelakowska, R.M., P.
a1ne, G, A, Taylorand K,
R0+arrap'), Europe, J, Ca
ncer s 165713r1980: J. Scanlon, B., A. Morotsuno, R. Bertino, J.
, B, Hynos (K, J.

5canlon %B, A, Moroson %J,
R, Bertino and J, B, Hynes
), Mo1. Pharmacol, +16, 2
61 (1979); o, o, Bird, J, W, Beitkus, J, Clark (0, D, Bird %J, W,
Va+tkus and J, C1arke), Mo
1. Pharmacol, 6.573 (1970
):]. Recently, N-14-(N-[(2-amino-4-
hydroxy-6-quinazolinyl)methyl] procedure 2-
Inylamino]benzoyl)-L-glutamic acid (5,
8-Sode Other IQ-Zoro i? Lugil leaf e) is thymil-
was identified as a potent inhibitor of tosynthetase [T,
R, Noyons, A, H, Calvert, A, L, Tsuyatsukuman, S.

J. Brown, M. Jones, ° To, Ro) 1 Lap (''.

RlJones 1A, HlCalvert s A,
L Jackman %S, J.

Brown %M, Jones and R, Ha
rrap'), Europe.

J, Cancer, 17.11 (1981)]. This enzyme catalyzes the do nova synthesis of thymidine nucleotides required for DNA synthesis.

Synthesis of pyridoC2,3-d]pyrimidine series derivatives by W, J. Ilwyn, OoG, Raipari (W, J,
lrwln and D, G, Wlbberl
ay), Advan.

Heterocycle, Chem, IQ, 14
9 (1969), this review covers the literature up to the beginning of 1968. Although many methods are reported in this review, the main synthetic routes for obscene systems include cyclization of functional derivatives of 2-amine nicotinic acid with various reagents [e.g., R1, Robbins, G.
H. Hitchings (Ro, Roblins and
G, H, Hitchlngs'), J, Am,
Chem, 5OC1+ 77.2256 (19-55
)) and the reaction of a 4-aminopyrimidine derivative with a 1,3-radical compound or a masking derivative thereof [for example, S, S.

Burl part, B, F, Valenti (B, S, Hu
rlbertand B, F, Valenti)
, J.Med.Chem., 1 1.708
(1968)). Pyrido [2,3
-d] Virimison was synthesized by R, Hernesotei, F, Mancini, C, C, 7'Rice (R, Bernetti).
, F. Mancini and C.C., Pr1c.
e), J, Org, Chem,, 27.286
3 (1962'l and B, S., Baalbert, B.
F, Valenti (B, S.

Hurlbert and B.F. Valenti
), J, Med, Chem, +11.708 (1
968). 5-Okin (8H
)-pyridoC2,3-d] The method for producing pyriminone is as follows:
B, H, Rizkara, A, D, Plume (B, H, R
izkalla and A. D. Broom), J.
, Org, Chem ++ 37.3980 (19
72). This reference describes the following compounds:

From the above compound, N-(4-C(2,4-noamino5-oxo(8H)-pyrido[2,5-d]bivillin-6-yl)methylamine]pe/diyl)-thi-daltaminoic acid (
5-deaza-5-oxoaminopterin), i.e., a compound having the formula A. Suri Nivasan A. D. Bloom [^, 5rinivas
and A, D, Broom), J.
Org, Cham,, 45.3746 (198
0) has been reported. Furthermore, N-[4-[(2-7 mi/-4
(5H)Oki7-10-*lumylviride [2,5-d
]pyriminon-6-yl)methylamine]benzoyl 1
-1-glutamic acid (5-deaza-10-pormylfolic acid) is G.

In, Smith, W. T., Mueller, P. A., Benokovinok, S. J., B./Kopik (G., N., Sm1th.

W., T., Mueller, P., A., Ben.
kovic and S, J.

Benkovic l, Biochem
istry, 20, 1 24 1
[Reported by 1981NC. A method for producing 5-deazafolate is not described.

Inhibition of bacterial nohydrofolate reductase by pyrido[2,3-cl]pyriminone was demonstrated by the above-mentioned Advan,
Heterocycle, Chem, is described in Ref. Recently, the pyrido[2,3-dl bilimino-1TS conductor has been shown to be a potent lipophilic inhibitor of mammalian nohydrofolate reductase. Le, o, s.

Danoch, C, ^, Nicole (E, M, Grivsky
, S.

Lee, C.W., Sigel, D.S., Duc.
h and CaA, N1chol), J, Me
d, Chem, 23.327 (19801). This reference describes the following compounds. Other derivatives of this ring system have been evaluated for antihypertensive activity. Thus, L, R , Pennett [L, R,
Bennett L., J., Med, C.
hem,.

(Guru, 38211981) reported that the following compound lowers blood pressure in hypertensive rats.

Synthesis of 5-deazafolic acid involves D, T, no-st ID,
An Introdu of Chemistry and Biochemistry of Biriminono, Purine, Zoterinono
c-tion to the chemistry a
nd Biochemistry of Pyrimid
ines, Purines and Pteridi
nes l", Noyo/Willia/Dosano's (Joh
n Wiley and So, ns, Ltd-12
31f 19801. 2--f i no 6-formyl-5-deazazoterino-4(38)-
The synthesis of 5-deazafolic acid using as an intermediate a compound having the formula C, tenozol, Jr, , J,
A. Monogomery (C. Ternple, Jr., a.
nd J., A., Montgo-Meryl, l.
Synthesis of anticancer drug with VJT ability (Synthe-s+
s of Potential Anticancer
Cancer Chem
therapy National 5service
Center, 5outhern Re5earch
1nstltut@I, Prodare Reboot
(Progress Report) 8511'
2 (1966) and Prodare Reboot (Prog
Res Report ) 86.8-10 pages (19
67). The synthesis of 5-deazafolic acid by a condensation reaction involving triformylmethane has been described by C1P, Dissertat CC, P, Tser+g), Dijon-Teinyong Abstract Int, B (Dissertat
ion Ab-structs Int, B), 4
0.3752 (198[]).

The original paper of this abstract, c, p, tsenda (c,
P, Tseng l, Research in Heterocyclic Chemistry (5tu
clies in Hetrocyclic Chemi
stry), 171-185 (1979) is a moat, 5-
Preparation of deaza-2,4-noaminozoterino/-6-carpiquinaldehyde dimethyl acetal, a compound having the formula, synthesis of 5-deaza-6-formylgtyrin, and acetylation of this compound, 5-deaza-6- It is also described that attempts to convert formylgtyrin to 5-7''' azafolic acid were unsuccessful.
The production of 5-7J azaaminopterin via a longer reaction 1 involving on l is shown in E and F.

Elslag, J., Dagol (E, F, Elslag
er and J, Davoll), 〃dLectures in Heterocyc
lic Chemistry)”, 2.5-97
, S-119 to S-121 (1974).

5-deaza analoda of folic acid, N10-substituted 5- of folic acid
Deaza analhoda, 5-0-deaza analhoda of aminopterin, 5-deaza analog of N-substituted aminopterin, 5-deaza analhoda of noethyl ester of aminopterin suppresses the growth of human epidermoid carcinoma cells f@A2 and in experimental animals. active against leukemia.

5-tea + J'7suO-1'' of folic acid referred to herein,
The 5-deaza analog of N-substituted folic acid has the following structural formula (I); ■): (Structural formula (1) above)
, (II>, R is hydrogen, CH3 or CH3CH
24 or CH3CH2CH2 or CH22CHCH2
CH=CCH2). Compounds of formulas (1) and (b) in which R is other than hydrogen are '# compounds.

The present inventors have now discovered a new intermediate useful for producing compounds of formulas (1) and (It). These novel intermediates have the structural formula ([11, ([lAl, flllEl) :(
In the above formula (fill, (IIIA), (me), Y
is CHO or CH20H or CH2Br.
ZUCH20H-j is CH2Br).

Structural formula ■: By reductive alkylation and alkylation of p-aminobenzoyl-sinoalkyl glutamate with a compound of formula (III) and a compound of formula (IIIA), respectively, structural formula ■: (■) (In the above formulas (■l, [■), R is the same as defined above and R1 is a lower alkyl group, i.e. an alkyl group containing up to 6" carbon atoms). Similarly, the formula (11B) of a compound of formula (Vll)
Alkylation with a compound gives a compound having the structural formula C■A): (in the above formula (■A), R and R1 are the same as defined above). When R in the compound of formula (■l is other than hydrogen, Y in the compound of formula (III)
and Z FiCH28r in the compound of formula (111B)
It is preferable that

Compounds of formulas (■) and (■A) can be converted to compounds of formulas (II) and (1), respectively, by saponification. Furthermore, the compound of formula (■) is R.

Tratner, G, Elion, G, Hitching 1.0°7
Yare7kin (R, Tratn@r, G, Elon,
G, Hltchlngs.

and D, 5harefkln), J, Org.
, Chem, 29.2674 (1974), the 4-amine group can be hydrolyzed to compounds of formula (1) by treatment with base under harsher conditions. Alternatively, the compound of formula (II) (R=H) may be added to sodium hydroxide. By methylation with formaldehyde in the presence of robidoride, the formula (n)
A compound (R=CH3) is obtained. Formula (K) (R=C
Compounds of H5) can also be obtained by methylating compounds of formula (1) (R=H) with a formaldehyde-sodium cyanogolobidelide combination.

The compound of formula (Ill) (Y=CHO) has the structural formula (■)
:In compound ke, Structural formula (V): C1-1[c)4=-N+(CH3)2)3 3X
(V) (in L genus (V), Ru.

The synthesis of triformylmethane was carried out by Z, Arnold and J, Zem.
Icka), Co11. Czech, Chem
, Commun, 25.1318 (1960)
and 2.了-Nold (z, Arnold), Co
11. Czech.

Cherrb Commun,, 26.3051 (
19S1). Thus, in one method, the complex ((C1-13) 2N=
CHCt) decaCt- with bromoacetic acid to give a quaternary salt (probably formula (Vl'); treatment of this quaternary salt with aqueous potassium carbonate gives triformylmethane. Isolation of triformylmethane and purification is difficult, and the method described herein uses intermediate quaternary salts or hydrolyzed derivatives thereof.

In water under reflux, the compound of formula (V) and 2,4°6-triaminopyrimidine (■) are condensed to produce 2.4-noaminopyrido[2,3-d]bilimino/-6-carpoxy. Aldehyde ((In): Y=CHO) was obtained.

The structure of this compound is obtained by oxidizing the formyl group with alkaline potassium permanganate and decomposing the 4-amine group tl-7111 to form the known 2-amino-4(3H)oxopyride, as explained later in Example 8B. (2,3-d) It was confirmed by the method to obtain pyrimidine-6-cargenic acid (■) (R,
Bernetstei, F.

Mancini, C, C, Price (R, Bernettl,
F.

Manclnl ar+dC,C, Pr1ce), J
, Org, Cham, 27°2865 (196
2) *OM, A. Lehery, S., G., Kotztis, H., Tiekelmann (D., M.).

Mulvery, S., G., Cottis and
H, Teckelmann), J.

Org, Cherr. 29.2903 (1964)
]. The reference sample of formula (Vl) was E, Stark, E, Preitreia (E), as explained later in Example 8A.

5tark and E, Breitmaier),
Tetrahedron, 29°2209 (197
3) 2-amino-6-methyl-4 synthesized by the method described
The alkaline permanate of (3H) ochinoviride [2,3-d]pyriminone was prepared by potassium heptaate oxidation. 2
, 4-noaminopyrido[2,3-d]biriminone system, it has been established that 4-aminopyrido[2,3-d]biriminone readily undergoes alkaline water splitting [R, Tratner, G.

Eliono, G, Hituchi/Guzu, 0.7 Yarevkin (R,
Tratner, G., Elion, G., Hit
chlngs and D.

5narefkin), J, Org, Chem,
, 19.2674 (1964)).

Although the mechanism of the condensation reaction is unknown, it is likely that two of the formyl groups or latent formyl groups of formula (V) react with the enamide/moiety of 4-aminopyriminone to eliminate either water or trimethylamine. do not have. The initial reaction consists of the formation of a Schiff base by electrophilic attack of one formyl group or derivative group by either the 5-position or the 4-amino group of the birinone ring, followed by ring formation of the resulting monocyclic intermediate. production of the desired bicyclic alkaline system by oxidation. Reference J, Org, Chem, supra, in which Pr.
observed that pyrido[:2,3-d]pyrimino/ is easily produced from 4-aminopyriminone and malonaldehyde containing an electron-withdrawing group under mild conditions. ing. Compound (V,) can be considered as a malonaldehyde derivative substituted with an electron-withdrawing group.

/ 7) /' ” The compound of formula (Ill) (Y = CH20H) is
Formula (I) in N,N-trimethylacetamide (DMAC)
Ill is produced by reducing the compound (Y=CHO') with sodium borohydride.

The compound of formula (lll) (Y2CH2B, ) is DMA
It is prepared by treating a compound of formula (III) (Y=CH208) with nopromotriphenylphosphorane in C. [Jl,) 9 Ino 9-1'' 6A, Monkoume 11-(J, R, Piper and J, A
, Montgomery), J. Org.

Chem., 42.208 (1977)].

This means that the corresponding 2,4-bis(()lJ7ff-
derivatives of the formula (l
resulting in the production of the compound nA'). The formula ([[l A
) is the compound of Piper, Mo/Comerly (Pi) listed above.
The formula ([[l) (Y = CH2
8r). The formula ([lA
) compound or formula (11) (Y = CH28r)
p-aminobeno! Although it can be used for the reaction with noalkyl monoglutamate (V+), it is preferable to use a compound of formula C11A) which is generated in situ, as shown in Example 3B.

A compound of formula (mB) (Z = CH208), i.e. 2-amino-6-(hyProxymethyl)-4(31
1) Okinoviride C2,3=d':l pyriminone can be prepared with the formula (m) (Y = CH208) by hydrolyzing the 4-amine group of the compound 11f
I can do that. Formula (I[l B ) (Z = (, H
2-amino-6-(bromomethyl) of the compound 2O8)
-4(3H) oquinviride [2,3-dl virimison (
The conversion to (m B ): Z==CH2B, ) is
S, Srinivasan, A, D, Bloom (S, 5rl
nlvasan and A, D, Broom), J
, Org, Chem, 46.1777 (198
1) achieved by treatment with phosphorus tribromide according to the method described. Furthermore, formula (11B) (Z = CH28r
) compounds are J.A., Mo. Gomery, J.D., Rose, C., Temple, J.R.
J, A, Montgomery, J, 00Ro.
se, C, Temple.

'Chemistry and Biology of Pterinnon' by J.R. and J.R. Piper)
Biology of Pteridines)”
(W, Zofriderer (W.

Pfleichlerer (ed.), Walter P. Grugter (Walter de Grugter), Berlin, 1976, p.

4B5], the formula (III) (Y =
It can be produced by hydrolyzing the 4-amino group of a compound of CH2Br) with 48% hydrobromic acid.

p-C substituted amine) noalkyl benzoyl-glutamate (■) A, l-1, Calvert, T.

R, Noyons, A, L, Jackman, S, J, Brown, Ni, R, Harasoge (A, H, Calvert
, T. R. Jones.

A, L, Jackman % S, J,
Brown and K, R8Harrap)
Author: Advances in Tumor Prevention, Detection, and Characterization, Volume 5: Human Cancer, Its Characterization, and Treatment.
vention, detection an
dCharacterization.

Vol, 5: Human Cancer, It
s Characterization and Tre
atment) “[W+Davis, +R1/% Rapp, G, Statophorus (W, Davis %, R
, Harrapand G, 5tathopoulo
s), ed., Excerpta mezoica (Excerpta mezoica)
Medica), Amsterdam, 1980, I)-
272) and T.R. Noyones, A.H., Calvert, A.L., Jackman, S.J., Brown, M.
ones, A, H, Calvert SA, L, Ja
ckman %S, J+dayrown %M, J
one's and K, R, Harrap)
, Europ.

J, Cancer s 17.11 (1981).

Formula ([1) (Y=
CHO) and water to form p-aminobenzoyl
Diethyl glutamate ((Vi): R=H, R,=C
2H5)' by reductive alkylation to
Yield of 5-deazaminopterin diethyl ester: 32
obtained in chi. 5-Deadeaminopterin ((ll), R = +
) was obtained. Methylation of the latter compound was achieved by 1) treatment of formula (u) (R=H) with formaldehyde and sodium/ano-lono idlide in an aqueous solution of H6,4 in yields of 85% 5-Deazamethotrexate (C11), R=C1-13) was obtained. Formula (II
) The structure of (R=CH3) is pre-prepared by oxidation using alkaline permanganate (1) Lamb V, as explained later in Example 8C! 12! 2-amine produced
4(3H') oxinviride [2,3-dllll'' Mino/
This was confirmed by obtaining -6 to calde/acid (■), thus showing that methylation occurred without force of either the 4-amino group or the 10-amine group. Methylation of the 4-amine group was removed from consideration by alkaline hydrolysis of the 4-amino group to form 5-deaza-10-methylfolate ((1), R=CHs).

A preferred route for preparing formula (1) (R=H) is 5-deazaminopterin diethyl ester (■) in an aqueous sodium hydroxide solution at reflux temperature: R=H,
J = IC2H5), substitution of the 4-amino group as well as hydrolysis of the ester function occurred to yield 5-deazafolic acid (('I), R-H) in 79% yield. Methylation of the compound of formula (1) (R=I→) with formaldehyde and sodium cyanozolohydride produces 5-deaza-10-methylfolate ((1), R=CHs)2)'m in a yield of 84%. Although it was
This was identical to the product produced by alkaline hydrolysis of the compound of formula (n) (R=c+3). Formula ([) (R=H and CHs), Formula (■) (R:H and CHs), the structure of 5-deazaminopterin diethyl ester was confirmed by elemental analysis, 'H-N M R1 mass spectral data. It was done.

A compound of formula (mB) (Z=CH2B, ) and p-
Diethyl aminobenzoylglutamate ((■): R=
N-(4-
[(2-Amino-4(3H) oxinviride [2゜3-d
]Biriminon-6-yl)methylaminocobenzoyl)
-L-glutamate ((■A): R== H, R1=
C2H5)t- was obtained, which was oxidized and converted into a compound of the formula (R=H).

Compounds of formula (1) and (■) form pharmaceutically acceptable salts with both organic and inorganic acids.

Examples of suitable acids for ring formation are hydrochloric acid, sulfuric acid, phosphoric acid, acetic acid, citric acid, oxalic acid, malonic acid, salicylic acid, malic acid, fumaric acid, succinic acid, ascorbic acid, maleic acid, dimethane sulfonate. be. The salt is prepared by contacting the free base form with an equivalent amount of Shintaku's acid in the conventional manner. The free base form can be regenerated by treating the salt form with a base. For example, a cold aqueous solution of a base can be used. Cold aqueous solutions of KFi, sodium hydroxide, potassium carbonate, ammonia and sodium bicarbonate are suitable for this purpose. The free base forms differ somewhat from their respective salt forms in some physical properties, such as solubility in polar solvents, but otherwise the salts differ from their respective free base forms for purposes of this invention. Equivalent to base form.

Therapeutic compositions containing compounds of formulas (1) and (■) are useful for ameliorating and treating cancer diseases in mammals.

When the active ingredient of the therapeutic composition is administered in an amount ranging from about 59 to about 200 m9 per 14 body weight per day,
Inhibits tumor growth in transplanted mice. Preferred doses for optimal results are from about 5 to about 50 mfil per body weight per day, but from 350 mfil to about 6.5 mfil total for a subject weighing 704 mm. of the active compound is administered in a 24 hour period. This dose can be adjusted to obtain the optimal therapeutic effect. For example, the dosage may be administered in divided doses each day or the dosage may be proportionally reduced as indicated by the exigencies of the therapeutic situation. A crucial practical advantage is that the active compounds can be administered in any suitable manner, such as by the oral or intravenous or intramuscular or subcutaneous route.

For example, the active compound can be administered orally with an inert diluent or an assimilable edible carrier, or can be enclosed in a hard or soft shell gelatin capsule,
Alternatively, it can be compressed into tablets or added directly to food. For oral therapeutic administration, the active compound may be mixed with excipients and formulated into ingestible tablets, buccal tablets,
ccal) It can be used in the form of tablets, troches, capsules, elixirs, suspensions, tablets, wafers, etc. Such compositions and formulations have at least 3.
It must contain one (1) active compound. The hex fraction in compositions and formulations can, of course, vary, but can conveniently range from about 2 to about 60 inches by weight. The amount of active compound in such therapeutically useful compositions is such that a suitable dosage is obtained.

Preferred compositions or formulations according to the invention are prepared so that a unit dosage form for oral administration contains from about 5 to about 200 inches of active compound.

Tablets, troches, pills, capsules, etc. may also contain the following ingredients: Binders such as gum tragacanth, acacia, corn starch, gelatin; Excipients such as dicalcium phosphate; Disintegrants such as corn starch, potato starch, alginic acid, etc.; Sweeteners such as sugar, lactose, saccharin, or flavoring agents such as mint, oil of wintergreen, cherry flavor may be added. When the dosage unit form is a capsule, it can contain, in addition to the materials enumerated above, a liquid carrier.

Various other materials can be present, such as coatings or to modify the physical form of the dosage unit. For example, tablets, pills, and capsules may be coated with shellac and/or sugar.

A syrup or elixir may contain the active compound, sugar as a sweetening agent, methylparaben and norobil as preservatives, a dye and a flavor, such as cherry or orange flavor.

Of course, the materials used to make unit dosage forms must be pharmaceutically pure and substantially non-toxic in the amounts used. The active compound may also be used for sustained release (5us
tained-release) formulations and compositions.

The active compounds of the invention may be administered parenterally or intraperitoneally.
can also be administered. Solution of active compound as free base or pharmaceutically acceptable salt td, preferably -1 L, <
can be prepared with water mixed with a surfactant such as d-hydroxypropyl cellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, and mixtures thereof and in oils. Under normal conditions of storage and use, these formulations contain preservatives to prevent the growth of microorganisms.

Pharmaceutical forms suitable for injectable use include sterile solutions or suspensions and sterile powders for the extemporaneous preparation of sterile injectable solutions or suspensions. In all cases, the form must be sterile and fluid to the extent that easy syringability exists. Injectable preparations must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier may be a solvent or dispersion medium, such as water, ethanol, polyols such as glycerin, noropylene glycol, liquid polyethylene glycol, suitable mixtures thereof,
Contains vegetable oil. Proper fluidity can be maintained, for example, by the use of coatings such as lecithin, by maintaining the required particle size in the case of dispersions, and by the use of surfactants. Prevention of the action of microorganisms can be carried out by various antibacterial and antifungal agents, such as noyarapen, chlorobutanol, carbolic acid, undruvic acid, thimerosal, etc. In many cases, it will be preferable to include isotonic agents, such as sugars or sodium chloride. Prolonged absorption of injectable compositions may be achieved by the use of absorption delaying agents (e.g.
aluminum monostearate, gelatin) in the composition.

Sterile injectable solutions are prepared by incorporating the active compounds in the required amount in the appropriate solvent with various other ingredients enumerated above, as required, or after sterilization. Generally, dispersions are prepared by incorporating the sterilized active ingredient into a sterile vehicle that contains the basic dispersion medium and the required ingredients from those enumerated above. In the case of sterile powder for the preparation of sterile injection solutions.

Preferable 8111m! ! The method is vacuum drying and lyophilization in which powders of the active ingredient and additional desired ingredients are obtained from their spun and sterile filtered solutions.

As used herein, a "pharmaceutically acceptable carrier"
Any and all solvents, dispersion media, coatings, antibacterial agents, antifungal agents, isotonic agents, absorption delaying agents, etc. are included. The use of such media and agents for pharmaceutically active substances is known in the art. This invention contemplates their use in therapeutic compositions, except that conventional media or drugs are incompatible with the active ingredients of the invention. Supplementary active ingredients can also be incorporated into the compositions according to the invention.

It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. As used herein, unit dosage form means physically discrete units suitable as unit doses for the mammalian subject being treated, each unit calculated to produce the desired therapeutic effect. containing a predetermined amount of active substance, together with any required pharmaceutical carrier.

The specifications for the unit dosage form are based on (a) the unique properties of the active substance and the therapeutic effect to be achieved, and (b) the disease symptoms in which pre-systemic health is compromised as detailed herein. is dictated by and directly influenced by the technical limitations in the formulation of such active substances for the treatment of diseases in a subject having a disease.

As noted above, the principal active ingredient is combined in effective amounts with a suitable pharmaceutically acceptable carrier in unit dosage form for convenient and effective administration. For example, unit dosage forms may range from about 0.1 to
It can contain an amount of the primary active compound ranging from about 400 to about 1, preferably from about 1 to about 60. Expressed as a ratio, the active compound generally contains about 0.1 to about 400 tag/a
1 carrier. In the case of compositions containing supplementary active ingredients, dosages are determined with reference to the recommended dosage and administration method for that ingredient.

The following examples demonstrate the best way to carry out the invention.

Example 1 2.4-noamino-6-(calcaldehyde (ca
rboxaldeiyde ) ) pyrido(2,3
-d) Pyridine (Ill: '/=CH0) into N,N-dimethylformamide (11,0t, 150 mmol) being cooled and stirred in a water bath5. r-+ phosphorus 74ide (27,51n/!, 46°01.600 mmol)
was applied IJD for 15 minutes. After stirring for 1 hour at room temperature, the reaction mixture was diluted with bromo vinegar (1(13,97,1
00 mmol). The resulting solution fzr was protected in a chloride tube, heated at 92C for 10 hours, and evaporated to dryness in vacuo.

The layered oil (~60i) was dissolved in water (1000 ml) and the resulting solution was neutralized to pH 7 with 1 Na 50 thihydroxide. 2,4,6-triamino bili
After addition of 7JI (5,000 mmol, 40.0 mmol), the bath solution was refluxed for 5 hours and filtered hot through fluted P paper. The P liquid was cooled and the precipitated solid f P11! The mixture was washed with water and dried under vacuum over P2O5.

Yield 12.5sy (33chi). Mass Hardness Torr, m/
e 1139 (vr+-). HPLC (0, 1M
NH40Ac (pH5,6)-Ch130H (9:1
)) showed that the purity of this product was >86. Sample (20, Of#) was diluted with 0.1N HCt (
15 m/) and diluted with acetone (225 fflZ) to precipitate the impure compound of formula (I[) (Y=CHO). The yield of 911n9°Pti was evaporated to dryness under reduced pressure and the residue t' was vacuum dried over P2O5 to yield the compound (■
) (Y=CHO) was obtained.

Yield, 128% omp: The color gradually darkened and decomposed with white sublimate at 360°C. λmaxnm (t
x 10-3) 0, I N -25 in HCl
8 (16,4), 317 (9,12), 3265h (8
, 24): pH 7-263 (15, 0), 316 (1
0,1), 345(10,8):0. IN NaOH - 254 (13,2), 267 (13,5), 316
(8,56), 374 (10,0). H-NMR(C
F3CO2D, 6 fb w/v), 964
88%9.75s(5-CH,7-CH), 10.21
S(6-cHo).

Analysis CB87N50"HCl" Calculated value as 1.3H20: C538,57:H,4,50:N,28.
12° Actual value: C, 38, 44; H, 4, 15: N.

28.14.

Example 2 2.4-noamino-6-(hydroquinemethyl)pyrido [
2,3-d]pyriminone ((■);”/=Cl-120
)-1) 2.4-noa e-noviride (2,3-d]pyriminone-6
-Cal, Gixaldehyde hydrochloride (2001%l, 0.
884 mmol) in DMAC (40 ml) solution, O
In CK, I N NaOH (0,8841 nl, 0
．． 884 mmol) and then treated with NaBH4 (1
DMAC (i + n
l) Treated with medium@turbid solution. The solution was stirred at 25° C. for 30 minutes and evaporated to dryness in a high vacuum knife. Residual 'fjlJ
k in H2O (5 ml) and, with stirring, adjust the pH to 3 with 6 N HCl and stir for 5 min.
The pH was adjusted to 7 with 1N NaOH. The precipitate was collected by filtration, washed with H2O, and dried under vacuum (P2O5). Yield 121 +Q (51%). Mass spectrum, ml
e191(+-)-t, 87 (M)+ (by DMAC), 36 (M)+ (by HCl-). λmax
nm (εX1O-5): 0, -274 in INHC6 (
6゜91), 318 (8゜21), 330sh (7,1
7), 362 (1,77): pH 7-248 (18,1
), 271 (9, 85), 338 (6, 52): 0,
IN NaOH-248(19,7), 271(
10,3), 345(7,14).

Analysis C3H9N5o・1.1Hct@o. 4 (CH3
) Calculated value as 2NCOCH3: c, 43.32:t
-+, 5.19:N.

28° 42° Actual measurement: C, 43, 48: H, 5, 2
4:N, 28. 48゜Example 3 N-[4-((2,4-diaminopyrido[2゜3-d)
pyrimidin-6-yl)methylamine]be/diyl)-
Diethyl L-glutamate ((■): R=H%R1==
c2+5) A, Dissolved compound m ('/=CH0) (1,47P, 5.90 mmol) in 70% warm vinegar (59 ml) and cooled to 25 C, p-aminobenzoyl-L
-1”/L/Tami7 acid) engineering ful (2, 28?, 7
．． Raney nickel (6,3
y, wet type! ! :) in the presence of 25C1 at normal pressure, 17
Hydrogenated for hours. The mixture was filtered and the catalyst was washed with 7o thiacetic acid (25wε). The filtration and washings were combined, evaporated to dryness at high Jc9F, and the residue was bathed in ethanol tif:
It was poured into 2N Na2CO3 (60ml).

This mixture? The mixture was stirred to obtain a homogeneous powder, which was drained, washed with water, and dried. The resulting powder was boiled in ethanol (
The molten gf dissolved in 415 raJ) was poured hot and evaporated to dryness in vacuo. Remain? l to CHCl3 (85tn
The mixture was crushed with e), collected by filtration, and the solid was washed with additional JD CHCl3 (40 ml). A suspension of this solid in ethanol (140 ml) was stirred for 20 minutes and frozen. The product was quenched by filtration and dried with JE (P2O5). Yield 945=n
9 (32%). mp, 262°C [Kofler Helzbank). Mass spectrum, m/e 496 (M +1).”m
axnm (gXlo-5): Q, IN HC6-2
18 (42,4), 280sh (19,3), 301J
(22,0): pH7-218(36,4), 249(
20,2), 280sh (22,3), 297 (23°6), 355sh (6,10): 0°IN NaO
In H-249 (22,0), 280 (23,8), 29
7sh (22,5), 645 (7,23). IH-NM
R (DMSO-δ6,6 excellent w/v).

δ1゜18 m (CH3), 2゜05 m (C, l
:! 2CH2CO)%2, 45t (CH2CO),
4, 08 m (CH2O), 4゜52 m (C
H2N, CHN), 6. 51 s, 7.51
s(NH2), 6°67d, 7.69d(C6H4),
6, 71 s (CH2N, h ), 8.25 d (
CONH), 8, 41 c! (5-CH, J=2.
0l-1z), 8.66d (7-CH, J=2.
0Hz).

Analysis Calculated value as C24H29N705: C158
°17; H, 5 °90; N, 19.79 ° Actual value: c.

57.91:H, 6,24:N, 19.55° The lower purity product (mp 246°C
) was obtained.

B. A solution of triphenylphosphine (430~, 1.64 mmol) in anhydrous DMAC (4rnl) was incubated at 0 °C.
It was then treated with bromine (84 μz, i, 64 mmol) under N2. The solution was stirred for 15 minutes at 00°C, treated with the compound obtained in Example 2 (100~, 0.0 mmol), stirred for 17 hours at 25°C, and was stirred with the formula (I[lA
) was obtained. This solution was treated with diethyl Thp-aminobenzoyl-thi-glutamate (194~, 0.603 mmol) and stirred at 25C for 17 hours, followed by ice water (4
0 ml). Obtained solution (pH2)
filtering to remove triphenylphosphine oquinide;
, adjusted to 1) 86.8 with 50% NaOH and cooled in a water bath. ■Product ((■): R=H, R1=C2H5
) was separated by centrifugation, washed with water, and dried under vacuum (P2O5). Yield 136++1 (73%). Retention time (HPLC) and Rf value (TL
C) was the same as that obtained from the analytical sample obtained in A above of this Example. mass spectrum, mle
496(M-to1)+.

Example 4 Methylamino]benzoyl)-L-glutamic acid ((1
): R=H) Product obtained in the previous example (■) (R=H, R, =C5H5
) (100 rv10.202 mmol) wo anoxic I
The suspension in N NaOH (4 iJ) was
Stir at reflux temperature for 4.25 hours under nitrogen, then dilute with 6N HC.
The mixture was acidified to pH 3.1 at t. The whole precipitate was dried for a week and dried under vacuum. This solid was dissolved in 1NHC6 (0°5ml)
The solution was diluted with water (Q, 5+n/), filtered, diluted with water (9+++/) and adjusted to pH 3 with 1N NaOH.
, 1. The precipitate was collected by filtration, PH3,
It was washed with 1 water and vacuum dried (P2O3). Yield 74~
(79%). mp Undefined. Mass spectrum, m/
e 441(M +1 )+. λ nmma× (gxlO'): 0. IN Hct-213 (37
,0), 280 (23,9), 2975h (20,6)
, 350 (7,55): pH 7-216 (40°8),
278 (24,9), 295sh (23,8): 0. I
N in NaOH-243 (22,9), 27B (24
, [3), 2qssh (22,7ha345 sh (7
゜58).

1HNMR (CF3CO2D - (6% w/v), 2
．． 56 (CH2C82CO), 2.82t (c+2o)
, 5.11m (CHN% C82N), 7.87a
, 8.15d (C6H5), 8.98s, 9°10S
(5-CH.

7-CH).

Analysis C20H20N606” Calculated value as 1°1H2o: C552,20:H,4,86:N, 18
．． 26° actual measurement unit: c, 52. 00:
H,4,92:N, 18° 54° Practical value 5 N-(4-[(2,4-noaminopyrido [2°3-d)
birimino-6-yl) methylamino[be//yl]-
Monoglutamic acid ((n): R=H) Product obtained in Example 3, (■) (R=H, R1=C2H
5) A solution of (359 m9, 0.724 mmol) in nomethylsulfoquinde (10 mJ) was diluted with IN NaOH (10 mJ) under N2.
1,81rnl.

1,81 mlJ%), stirred in a stoppered flask under N2 for 6 hours, and evaporated to dryness in vacuo at <jO'c. A solution of the residue in water (18 ml) was filtered and acidified to pH 3.6 with INHC6. The precipitate was filtered off, washed with water at pH 5.6, and dried in vacuo (P2O3). Yield 297η (87%). mp undefined (softens above 20d℃).

Mass spectrum, m/e 440 (M+1)+.

λmax nm ('X10') ” 0
- IN HCZ -218 (40°5), 280
sh(16,9), 300(18,8): pH7-21
8 (38,5), 245 (19,2), 280 (23,
9), 296sh (22,7): 0-IN Na
In OH - 2'48 (22,0), 280 (24,4),
296th (22,7), 545(7,75).

IH-NMR (oMso-H6, 6% w/v), 22.
00m(CH2Cl-12Co), 2.29t(
CH2Co), 4.56 m (CHN, CH2N
), 6.66d, 7.68d (C6H4), 7°41
(NH2), 8.04m (NH2, NH, Co2H
), 8°52d, a, 70d (5-c) (.

7-CH).

Analysis C20H21N705" Calculated value as 1, 9 +20: C, SO. 72: H, 5, 28: N
, 20. 70° actual measurement 1st shift: C, 50, 86;
H, 5, 45:N.

20, 50° Example 6 -r Mino]pennoyl)-thi-glutamic acid ((II
):R=CH3) Compound (It) (R-H) (100 μm, 0.211 mmol) was suspended in oxygen-free water (5 μm), and a cell solution was prepared by adding 4 μm of cell solution. The pH was adjusted to 6.4 with 1N NaOH, and the solution was washed with gold 38% + cho (83.1μt, 1.5μt).
14 mlJ mol) and then NaBH3CN (
19.9 to 0.317 mmol). This solution was added slowly for 45 minutes or once with 1N HCl/10
The pH was maintained at 6.4 by The solution was heated with N2
Stir for 26 hours at
made acidic. The product (collected by filtration, pH 3
Wash with water at °6 and vacuum dry (P2O5) 1. Ta. Yield 97IR9 (94 chi).

mp undefined (softens and darkens above 217°C).

Mass spectonope m/e 454 (M + 1
)+.

λmaxnm (gXlo'): 0. IHC
-221 (37,1), 311 (19,0) in t:
pH7-219 (35°1), 247@(18,1),
305(25,2): 0-IN NaOH-249(19,9), 305(25,0), 355s
h(6,15).

IHNMR (DMSOd6, (5'A w /su, 62
．． 00m (Cs2CH2Co), 2.28t (CH2C
o), 5.12s (CH5), 4.32m (CHN),
4.66s (CH2N), 6.78d, 7.72d
(C6H4), 8.31d(5-CH), 8.59d(
7-c+).

Analysis C21H23N905" Calculated value as 2H20: c251°53; H, 5,56:N, 20.03.
Actual value: C, 51°54; H, 5,47; N.

20゜ 35゜Example 7 -N-(4-C[(2-amino-4(3H)-oxy7
Bi! J do[2,3-d]pyriminone-6-inob)+
１１ -１ -咄 -ト１ □
-Methyl]methylamino]benzoyl)-L-glutamic acid ((1): R=CH3) A6 formula (+) (R=H) (60~, 0.13 mmol)
g of a compound of formula (It) (R=CH,)
was methylated using the method used to prepare the compound. After the reaction solution was diluted with oxygen-free water (5 ml), INH
It was acidified to pH 3.1 with C4. Enjoy the product, pH 3
, washed with 1 water and vacuum dried (P2O5). Yield 53
m9 (84%). mp undefined. Chastity spectrum, m/e4
55 (M+1 Tsuji.

λmax nm (ε X1O-3) 20,
INHCt -215 (35,1), 280 (19
, 0), 306 (20, 8), 355 sh (6, 8
5): pH7-216 (38,0), 274 (19
, 0), 306 (27°6); 0°lNN-2 in NaOH
42(22,9), 275sh(17,4), 307(
25,4). IH-NMR (DMSO-d, 5
CH w / v ), δ2 , 02 m (CH,, C
H2Co2H), 2°35t(CH2CO), 3.
09s (CH3), 4.37m (CHN), 4,
73s (CH2N), 6.82d.

7.75c+(c6+4), 8.03d(5-CI-1
), 8°19 d(NH), 8.55a (7-c+).

Analysis C21H22N605・H20・o, 75Hct
Calculated value of toshite: C, 52, 14: H, 5, 16: N%
17°37° Actual value: C, 52, 12:H.

5, 12:N, 17. 47°B, compound e+(II) (R=CH5) (50q)
o solution was hydrolyzed by the method of Example 4 for the preparation of compound (1) (R=H) to obtain compound (+) with a yield of 64 yen.
(R=CH3) was obtained. HPLC and UV data are
This product was shown to be identical to that prepared above.

Example 8 2-amino-4(3H)-oxopyride [2°A, I
N NaOH (2-amino-6-
While stirring a solution of methyl-4(3H)-oxopyrido[2,3-d)pyridine (177m9.1.00<limole) at reflux temperature, about 1 mol of 0.2M potassium permanganate aqueous solution was added. I spoke to him over time. After decomposition with excess potassium/sodium dihydrogen formate, the resulting hot mixture was filtered through Celite. The P solution was adjusted to pH ~3 with HCl and left at room temperature for 18 hours. The precipitated solid (~170) was removed by filtration, dissolved in 2N NaOH, and the bath was cooled to precipitate the sodium salt of the product. The salt was collected (collected by filtration, dissolved in water, and the resulting solution was purified with HCt).
Adjusted to H2-6. All of the precipitated solid was filtered and dried under vacuum over P2O5. Yield671)y
(29%). m p 265 Co H
PLC[0, IM Na2HPO4 (pH 7)
-CH3CN (9228)] indicated that the sample was homogeneous. 'max nm(εx10-
5):0. IN HCt-216 (35,9X266
(14,5), 306 (6°70), 515sn (5,
35): pH 7-216 (26,8), 232sh (1
7,8), 283 (11,4), 310sh (5,93
), 321sn(5,37); 0, IN NaOH-2
46 (22,8), 292 (10,1), 332 (7,
20). IH-NMR (Naoo, 5%w/
v ), δB, 76d (7-CH.

'~1.5Hz), 9.06d(5-CH).

Analysis Calculated value as C3H6N4o3-0, 6 HCt: C, 42, 13: H, 2°92;, N, 24
．． 57゜Actual value: C, 42゜04:H, 2,80:N
.

24° and 41° days, 2,4-noaminopyrido C2,
3-d) Virimidine-6-carmexalf)'((
II): Y=CHO) (186#I9, ~0.980 mmol) resulted in hydrolysis of the 4-amino group and oxidation of the polmyl group to yield the compound of formula (■). Yield: 158 Da. Field desorption mass spectrum, m/·206 (M+). HPLC chromatogram of this product (co-inject, lon)
) was the same chromatogram as the compound (■) obtained in A above.

C, N-(4-[[:(2°4-noaminopyrido(2,3-d)pyrimino/-6-yl)methyl]methylamino]benzoyl)-L-glu p in 2 ml of INNa□H
Minic acid (((1); R=CH3) (5,00■, 0.0
KMnO4 (1°62
9, a, a1as remol) and 950 for 6 hours 7
II] Heat, filter and adjust to 9H8 with IN HCt. The HPLC chromatogram shows the presence of the compound of formula (■) (yield ~22%) and the unreacted compound of formula (b) (day-CH5) (yield ~50%)? Indicated. The ultraviolet spectrum of the eluted compound (Vl) was the same as that of the reference sample of compound (■) in the ultraviolet light kettle.

Compound (1), (It), (■) (R=H, R1=C2
H5) cell culture cytotoxicity and mouse phosphorus
The activity against cell leukemia P388 is shown in Table 1.

1 Table 11-'- Cell culture cytotoxicity data 8 and activity against murine phospho-9 cell leukemia P388 5P388d Compound ED 0μM0 Dosage 1~/
ILsI fR=H) 6.1
100 171 [R=CH3111,2200
111t fR=H) 0.013 −
11 (R=CI-1310,004492@■(R=
H, R1=C2H510,0521118me) )l/
Kise) 0.001 2 61
'a) Human epidermoid carcinoma cells A2R,1,ke' Ruff, N,H.

Greenpelg, M.M., McDonald, A.M.

/ Neematsuha 1 Skin”, Azat (R, 1, Geran
.

N.H., Greenberg, M.M., MacDo
nald, A.M.

Shumacher, and B.J. ^bbott
), Cancer Chemotherapy Reports (Cancer
Chemotherapy Reports),
ρart 3. Vol, 3 (JEL2), 197
1°b) See al. C) Concentration that inhibits colony formation by 50% after 12 days, determined in ml in a plastic flask. L.

Shi, be 4 throat, "2+ H, D, '7 Yuinotei, N
,H,4il,P,W,ara/,J,A,monogomery (L,l,.

Bennett, Jr., H.D., 5chnet
ti, N, H+ Vail, P.

W, A11an, and J+A, Montgom
ery l, Mol.

Pharmacol, 2.432 (1966).

d) Inoculated intraluminally with 10 P38 am cysts; tcDr1 mice. The four products were administered intraperitoneally every day for 1 to 5 days.

el　One animal survived on the 30th day of Vi.

f) Ishiguchi, intraperitoneal administration for 1 to 9 days.

Claims (1)

[Claims] (1) In the structural formula C above, Y ViCHO and CH20H are CH2Br, and 2 is CH20H or CH2Br.
) for the production of a 5-deaza analog of folic acid, a 5-deaza analog of N10-substituted folic acid, a 5-deaza analog of aminopterin, a 5-deaza analog of N10-substituted aminopterin, Useful intermediate. (2) The intermediate according to claim (1), wherein the intermediate compound is 2,4-noamino-6-(calgaxaldehyde)pyrido[2,5-d]pyriminone. (3) The intermediate according to claim (1), wherein the intermediate compound is 2,4-diamine-6-(hydroxymethyl)pyridoC2,3-a)pyrimidine. (4) Formula: (In the above formula, R is CH3 or CH3CH2 is CH
3C82CH2 or CH2= CHCH2 or CH
of N10-substituted folic acid, characterized in that it has
-Dea the Ana Robe. (5) Formula: (In the above formula, Rij CH3 or CH3CH2 or CH3CH2CH24 Tij CH2= CHCH24
A 5-deaza analog of N10-substituted aminopterin, characterized in that it has the following Jd CH=CCH2. (6) The compound according to claim (4), wherein R is methyl. (7) The compound according to claim (5), wherein R is methyl. (8) % The compound according to claim (1) has the formula:
(-In the above formula, R is CH3-j or CH3CH2 or CH3CH2CH24 or CH2=CHCH2 or CH=CCH2, and R1 is a lower alkyl group)
folic acid or N10-substituted folic acid or aminopterin or N1
A method for producing a sialkyl ester of a 5-deaza analog of a 0-substituted aminopterin. (9) The method according to claim (8), wherein R1 is ethyl and R is hydrogen or methyl. tll R is other than hydrogen and Y group and 2 groups are C
Claims characterized in that it is H28r,
8) The method described in section 8). of the compound according to claim (7), which comprises methylating 5-deazaaminozoteri/ by reaction with formaldehyde in the presence of 11-sodium cyanolobidolide. Production method. +121 A process for producing a compound according to claim 6, characterized in that it comprises methylating 5-deazafolic acid by reaction with formaldehyde in the presence of sodium cyanoporohydride. (13) The compound according to claim (6), which comprises treating 5-deazamethotrexate with a base to hydrolyze the 4-amine group of 5-deazamethotrexate. Production method.