CA1196914A - Anthracycline derivatives and aglycone intermediates thereof - Google Patents

Abstract

ABSTRACT OF THE DISCLOSURE

New 4-demethoxy-11-deoxydaunomycin and 4-demethoxy-11-deoxyadriamycin anthracycline derivatives have been prepared and found to be useful antimicrobial and antitumor agents.
Further, intermediates, having the formula:

Description

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$~-9?89 BACKGROUND OF THE INVENTION
.

1. Field of the Invention This invention relates to new anthracycline deriva-tives, to methods for their use as antimicrobial and anti-tumor agents, to pharmaceu*ical compositions containing them and to synthetic methods and intermediates for the preparation of said new derivatives.

2. Description of the Prior Art Daunomycin (see UOS. Patent 3,616,242) and adriamycin (see U.S~ Patent 3,590,028) obtained from fermentation broths Of microorganisms of the genus Actinomyces are known as anthra-cycline-type antibiotics. These compounds have a broad anti-tumor spectrum against experimental tumors and are widely used clinically as chemotherapeutic antitumor agents.
Despite the usefulness of daunomycin and adriamycin, there is still a need for new anthracycline derivatives which will have greater antitumor activity and/or reduced toxicities~
To satisfy this need, attempts have been made to provide anthra-cycline derivatives by fermentation, biotransformation and both semisynthetic and total sy~thetic processes, Illustrative of these attempts are the methods di~closed by F~ Arcamone in Topics in Anti~ioti~ Che~i~ry ~:lQ2-279 (1978) and in U,S.
Patent 3,5~8,315 ~aclacinomycins A and ~).
U.S. Patent 4,247,545 disc~oses the preparation of 11-deoxy derivatives of daunomycin and adriamycin (as well as their respective aglycones) by fermentation of ~ peucetius var~ caesius (ATCC 31365).

~L96~

SUMMA:RY OF HE INVENTION

This invention relates to novel anthracycline glycoside antibiotics having the formula O

O OH O

~:3C ~

HO

wherein Rl is hydrogen, hydroxyl or (lower)alkanoyloxy, and nontoxic acid addition salts thereof, and to their respective aglycones having the formula O O

O OH OH II

wherein Rl is as defined above. The compounds included within the scope of formula I exhibit both antimicrobial and antitumor activity. The formula II aglycone compounds are important intermed;ates i~ preparation of the anthracycline end-products of formula I. Compounds of formula I in which Rl is (lower)-alkanoyloxy are also intermediates in preparation of the formula I products in which Rl is hydroxy. In tests with experimental animal tumors, the preferred formula I compounds (i.e. those in which R i~ H or OH) display espPcially s~rong antitumor activity.

~6~4 As used herein and in the claims the term "non-toxic acid addition salt" is meant to include all those organic and inorganic acid salts of the compounds of formula I which are conventionally used as substantially nontoxic salts of medicinal agents containing amine functions. Illustra~
tive examples would be those salts formed from such pharmaceu-tically acceptable acids as hydrochloric, hydrobromic, sul-~uric, phosphoric, acetic, propionic, maleic, oleic, palmitic, citric, succinic, tartaric, fumaric, glutamic, pantothenic, laurylsulfonic, methanesulfonic and naphthalenesulfonic. The term "(lower)alkyl" as used herein includes both straight and branched chain saturated aliphatic hydrocarbon radicals having from 1-6, preferably 1-4, carbon atoms inclusive, e.g. methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, n-hexyl, etc. Similarly, the term "(lower)alkanoyloxy group" denotes a residue of an alkanoic acid whose alkyl moiety is linear or branched and contains 1-5, preferably 1-3, carbon atoms. Examples of such groups include acetoxy, propionyloxy, n-butyryloxy, isobutyryloxy, n-valeryloxy and isovaleryloxy. The acetoxy group is an especially preferred alkanoyloxy group.
The compounds of the present invention may exist as the individual diastereomers or as mixtures of such isomers.
It is specifically intended that the inventisn include the resolved diastereomers as well as mixtures of such diastereomers within its scope.

Thus the present divisional specification provides a pro-cess for producing an aglycone intermediate of the formula C,'~ 2-OH

~.
. .

- 4a -wherein Rl represents hydrogen, hydroxyl or (lower) -alkanoyloxy, which comprises the steps of A) subjecting a compound o~ the formula O Y

O o~ W
o wherein Y represents -C-CH2-(lower)alkanoyloxy or ~z/o -C-CH3 in which Z is a ketal residue and W represents a halogen atom, to hydrolysis;

or where the intermediate of the foxmula ~ ~ CH3 is desired; the alternative step of B) reactiny a comPound of the formula.
~H 3 with phthalic anhydride of the formula ~b in the presence of a Lewis acid and in the absence of a solvent at a temperature of at least 150 C~
DETAILED DESC_IPTION
The c~mpounds of the present invention may be prepared by total synthesis from 5-methoxy-2~tetralone, a known compound, by the synthetic route shown in the following reaction scheme.

6~

REACTION SCHEME
-S

-- CH ~-CH3 (1) (2) (3) O H
(4) ~H3 O OH
R COOM
O O (8) ~ / I~ 1l 3 o C-CH20CR

~H

~.1' , O C~ ~ 3 ~ ~
~C-C~R ~ _ HCH3 ~ X2 0 0}~ X
(7) ~9) 2 ~- .~
~J

H _Rl2 H ~ a) 0~

y NHR~
R"' O

~ /

O ll CH R12 ~H
O'f~

H3 ~J
~/ NHR "
R"' O
(I-a) 0~ p O J
H3C1~
r NH2 OH (I~

~9~4 In the above reaction scheme, Rl is as defin~d above, R12 represents a hydrogen atom or a (lower)alkanoyloxy group, R3 represen~s 2 (lower)alkyl group, Xl and X2 represent a halogen atom ~Cl, Br, F, I), especially a bromine atom, Z
represents a ketal residue, M represen~s an alkali metal atom, an alkaline earth metal atom or an ammonium group, and R" and R"~ respectively represent a conventional amino-protecting group and a conventional hydroxyl-protecting group which can be easily eliminated by hydrolysis.
Each of the individual steps shown in the above reaction scheme may be practiced by known methods. The reactions in the individual steps are further elaborated on below.

~_==~

In this step, 5-methoxy-2-tetralone of formula (1) is reacted ~ith a Grignard reagent of the for-mula (CH_C)MgX3 wherein X3 represents a halogen atom, preferably a bromine atom, to yield a tetraline derivative (2). This reaction can be carried out by utilizing a known Grignard reaction, for example as shown below in Step A of Example 1.

(2) ~ 3) In this step, hydrolysis of the tetraline derivative (2~ gives 2-acetyl-2-hydroxy-5~methDxy-1,2,3,4-tetrahydro~
naphthalene. ~he hydrolysis can be carried out, ~or example, by treating c~p~und (2) ~ith suluric acid in the presence of mercuric oxide, mo~t advantageous7y at room temperatureO
The resulting compound (3) can be isolated from the xeaction mixture by a conventional purification procedure, for example chromatography.

69~
' - B -

(3) ~ (4) In this step compound (3) is reacted with phthalic anhydride to give 4-demethoxy-7,11-dideoxydaunomycinone (4).
The Friedel-Crafts acylation may be conducted according to the general procedure disclosed in Experientia 34:1255-1257 ~1978). For example, the reaction can be carried out in the presence of a Lewis acid such as aluminum chloride, titanium tetrachloride or zinc chloride at a temperature of from about 50~ to 250C, preferably 150-200C., for a period of from about 1 to 30 m-nutes, preferably about 5 to 10 minutes. The phthalic anhydride is used in an amount of at least one mole, preferably 1.5 ~o 2 moles, per mole of compound (3). The Lewis acid is used in an amount of at least 8 equivalents, preferably 12 to 20 equivalents, per mole of compound (3).
Most pre~erably, the Friedel-Crafts acylation is carried out in the a~sence of a solvent at a relatively high temperature at which the starting materials substantially melt, i.e. at least above 150C and preferably 170 to 190~CD~ This effective ly prevents the occurrencé of the side reaction mentioned below.
If the Friedel-Crafts reaction is carried out in the presence of a conventional organic solvent or a~ a relatively low temperature (less than about 140C), a compound of the formula ~-CH3 OH

. ~ OH

forms as a by-product in additi~n tn the desired oompound (4) and decreases tne yield of (4). It has been found that if the reaction is carried out in ~he absence o~ solvent and at a relatively high temperature, the compound (4) can be obtain-ed ~electively in high yield without the formation of undesired _ 9 _ by-product.

~4) , (5) ~ (6) This reaction can be carried out in a manner known ~ se, for example by the method describPd in U.S, Patent 3,803,124. Alternatively, the halogenation of compound (4) can be carried out by using an N-haloacid imide such as ~-bromosuccinimide or N-chlorosuccinimide, preferably N-bromosuccinimide, resulting in selective halogenation of the 14-position of compound (4), Halogenation with the N-haloacid imide can be carried out at a temperature of from about 0 to 50C. for a period of from about 2 to 10 hours.
The amount of the N-haloacid imide used is not critical;
advantageously, it is employed in an amount of at least 2 moles, preferably 8 to 16 moles, per mole of compound (4)~
The acylation of compound (5) with a salt of an organic carboxylic acid having the formula R COOM can be carried out at a temperature of from about 0 to 80C, for a period ffl from about 5 to 20 hours, Preferably there is employed an inert solvent such as a ketone (e.g. acetone or methyl isobutyl ketone) or an ether (e,g. tetrahydrofuran or dioxane) either alone ox as a mixture, The acylating agent is generally used in an amount of at least 1.5 moles, p~efer ably 2 ~o 4 moles~ per mole of compound ~5)~ Examples of suitabl~ acylating ag~nts i~clude sodium acetate, potassium aceta~e~ ammoniu~ acetate, sodi~m propio~ate, potassium pro-pionate, amm~ni~ pr~pi~nate r ~odium valerate and ammonium valerate~

(4) ~ (8~

The reaction in this step is ketali~ation of the carbonyl group at the 13-position of compound (4) which can be carried out in a manner known per se using a carbonyl protecting reagent (kentlizing reagent) known per se. For expampe, ketalization can be performed by the action of a ketalizing agent of the formula Ho-Z_OH on compund (4) in a suitahe inert solbent in the presence of an acid catalyst.
for example an aaromatic sulfonic acid such as p-toluene-sulfonic acid or benxenesulfonic acidl. The reaction tempera-ture is not critical. Gernerally, it is preferred to use tempareture of from about room temperature to the reflux temperature of the reaction mixture, most preferable a temperature of from about 80 V. to reflux temperature. The ketalizing agent ios used in an amoutn of at least 1.5 moles, preferale from about 5 to 15 moles, per mole of compound (4).
Specific examples of the detal residue Z which may be employed are -CH2CH2-, Spedific exampled of suitable ketalizing agents include ethylene glycol, poropylene glycol, 1,1-dimethoxypropane and triethoxymethand.

(6) ? (7)and (8) ? (9) These steps involve halogenation of he 7-position of compound (6) or compound (8). The halogneation can be carried out by using known hlaogenating agents such as bromine, chlorine, N-bromoxuccinimide or N-chlorosuccinimide in a conventional manner. Bromination is especially preferre.
In a preferred embodiment, a solution containing bromine is added to compound (6) or (8) in an aqueous medium in the presence of a free radical generator such as 2,2-azobisisobutyronitrile (abbreviated AIBN) and the reaction is carries out at a teimperature int ehr range of about 0 to 60 C, ~19~

preferably at room temperature, for a period of from about 1 to 10 hours. Advantage~usly, at least 1.5 moles, prefer-ably 2 to 3 moles, o~ bromine is used per mole of compound (6) or (8).

(7) or (9) _ ~ (II a) The reaction in this step is hydrolysis. Since compounds (7) and (9) are unstable, when the above halogenation step is carried out in the presence of water, the hydrolysis of compound (7) or (9) readily occurs to give the corresponding compound of formula II-a. The above hydrolysis may be conduct-ed by a conventional method using an aqueous alkaline solution under mild conditions, for example at a relatively low tempera-ture of from about room temperature to about 50C. using a weak base such as sodium hydrogen carbonate.
When the hydrolysis of compound (7) is performed under relatively strong hydrolyzing conditions, for example by treating the compound with 10% K2CO3 at room temperature for one hour, a compound of formula II-a wherein R12 is a hydroxyl group is formed.
Compound II-a i5 generally obtained as a mixture of four stereoisomers having the configurations (7S,9S), (7R,9R~
(7R,9S) and (7S,9R). A racemic mixture (7S,9S; 7R,9R) and a racemic mixture (7R,9S; 7S,9R3 can be easily separated from the stereoisomeric mixture ~f compounds II-a in a known manner, for example by a chromatogxaphic technique.
It is exp~cially preferred that the desired end-produc~ of formula I have the configuration (7S,9S~. ~ccord-ingly, ~he separated racemic mixture (7R~9S; 7S,9R) of compou~d II-a can be epimerized to a racemic mixture (7S,9S; 7R,9~).
As a result, the yield of the latter desired mixture can be increased, The epimerization may be carried out, for example, by contacting the above-mentioned racemic mixture of compound `' - 12 -II-a with an inorganic acid such as hydrochloric acid or perchloric acid in a water-miscible organic solvent such as acetone, tetrahydrofuran or dioxane at a temperature of from about lO~C to the boiling point of the solvent.

(II-a)--~ (I-a) -In this step the aglycone compound of formula II-a is reacted with a glycal of the formula 0~

R"' ~ III
derived from daunosamine to give a glycoside of formula I-a.
The amino-protecting group R" in formula III is selected from conventional amino-protecting groups which are easily eliminated by hydrolysis. Specific examples include (lower~alkanoyl groups such as acetyl, propionyl and tri-fluoroacetyl, aromatic carbonyl groups such as benzoyl and p-nitrobenzoyl, aralkyloxycarbonyl groups such as benzyloxy-carbonyl and alkyloxy~arbonyl groups such as t-butoxycarbonyl.

The hydroxy-p~tecting groups R"' are also selected from conventional hydroxy protecting groups which are easily eliminated by hydrolysis and may be similar to the groups mentioned above fo~ the amino-protecting group R~' .
The glycosidation reaction can be carried out in a conventional manner using an acid catalyst~ Typically, it is carried out in an anhydrous inert organic solvent, e.g. benzene, toluene, tetrahydrofuran or dioxane, pre~erably in an aromatic hydrocarbon such as benzene or toluene, in the presence o~ an acid catalyst, for exa~ple an aromatic sulfonic acid such as p-toluenesulfoni~ acid or benzenesulfonic acid ~r an alkylsulfonic acid such as methanesulfonic acid or butanesulfonic acid The reaction tel:lperature is advantageously in thc range of about 0 to 8DDC, preferably a~out 20 to 40~C. It is preferred to use the compound of formula III in an amount of at least 1~5 moles, ~3~91~

preferably 2 to ~ moles, per mole of compound II-a.
The glycal of formula III can be ~roduced by treating a daunosamine derivative having the 3-~mino group and the 4-hydroxyl group protected as de~cribed above with a sulfonylating agent such as ~-toluenesulfonyl chloride or benzenesulfonyl chloride in the presence of a base such as pyridine, dimethylaniline, morpholine or triethylamine either in an organic solvent or in the absence of solvent (see Japanese Published Patent Application 27346/79.
The glycoside of formula I-a resulting from the above-described glycosidation reaction is usually obtained as a mixture of an isomer whose amino sugar residue has a 1' linkage (to ~e re~erred to as a 1'~ isomer) and an isomer whose amino sugar residue has a l'~-linkage (to be referred to as a 1'~ isomer) with the 1'~ isomer generally predominating.
Accordingly, when the racemic mixture (7S,9S; 7R,9R) of the compound II-a is used as a starting material, the glycoside of formula I-a is a mixture of our stereoisomers (7S,9S,l'~), (7S,9S,1'~), (7R,9R~l~a) and (7R,9R,l'~), These isomers may be separated by conventional procedures such as chromatography on silica gel or the like, Isolation of these stereoi~omers is preferably performed after the de protecting reaction to be described below.

(I-a ~ I) This step is the elimination of the amino and hydroxy-pr~tec~ing groups of compound I-a~ This deprotecting reaction is carried out by a conven~ional hydrolysis method~
For example, it can be effected by alkaline hydrolysis at a rela~iv~ly low temperature of from abou~ O~C ~o room tempera-ture in the presence of a base such as sodium carbonate or potassium carbonate, This hydrolysis results in the elimina-tion of the protective groups R" and ~'~' from compound I-a.

-When ~12 is a ~lower)alkanoyloxy group, the lower alkano~1 group is also elimina~ed depending upon th~ conditions of hydrolysis to give a compound of formula I in which Rl is OH.
By the above-described procedure, the anthracycline derivatives of formulae I and II may be obtained in good yield.
In a variation of the above procedure, a compound of formula I-a in which R12 represents hydrogen can be con verted to the corresponding compound of formula I-a in which R12 is a (lower)alkanoyloxy group by sub~ecting it to the steps (4), (5) and (6) in the above-described sequence, Products of formula I obtained in the above reaction procedures may be recovered in the form of the free base, an acid addition salt or a nontoxic-acid addition salt. The free base products may be easily converted into nontoxic acid addition salts which are substantially equivalent in therapeu-tic activity to the corresponding free bases. The salts are formed, isolated, purified and formulated by the methods gPnerally ~mployed in salt formation for the anthracycline glycoside antibiotics, Thus, the free base may be reacted with a nontoxic organic or inorganic acid in a sui~able solvent and the salt recoYered by lyophilization or by precipi-tation with an antisolvent! i.e.~ a sol~ent in which the desired salt is only slightly insoluble. Products in the form of an acid addition salt may be converted to the corres-ponding free base by neu~ralization with a basic substance.

Finally, toxic acid addition salts may be conver~ed to non-toxic acid addition sa~ts ~y neutralization and treatment with a nont~xic acid as descri~ed ahove.
The aglycone moiety shown in the compounds of the present invention is drawn in a planar structure, but is meant to include all configurations (7Sl9S)~ ~7R,9R), ~7S,9R~ and (7~,9S).

9~

Biol~ical Properties The compounds of formula I, especially those in which R is hydrogen or hydroxyl and nontoxic acid addition salts th~reof, have sh~wn excellent antitumor activity against L1210 leukemia cells in culture and experimental animal tumors.
Especially preferred are those having the configuration (7S,9S) which is the same as daunomycin or adriamycin produced by fermentation. ~he glycoside moiety in the compounds of the present invention most preferably has a l'a-linkage.
The antitumor activity of the compounds of formula I
may be demonstrated by the following experiments.

A. Inhibitory effect on growth, ~NA synthesis and RNA synthesis of L1210 cultivated leukemic cells of mice The compounds of formul~ I markedly inhibit the growth and nucleic acid synthesis of cultivated leukemic L1210 cells of mice. For example, L1210 cells were inoculated in a concentration vf 5xlO cells/ml in an RPMI 1640 culture medium (Roswell Park Mem~rial Institute 1640) containing 20~
calf serum and, simultaneously, representatiYe compounds of the present invention were added in a concentration of 0.1 and 0.5 ~g/mlO The cells were cultivated in a Co2 generator at 37C. The 50% growth inhibitory concentration with respect to a control group was then determined.
Separately, the above L1210 culti~ated cells were suspended in a ccnce~tration of 5x105 cells/ml in an RPMI
1640 medium containing 10% calf serum and cultivated for 1 to 2 hGurs in ~ CO2 incubator at 37C. Then, representative test compounds were added in various concentrations, and 15 minutes later, 14C-uridine (0,05 ~Ci/ml) or 14C-thymid.ine (0.05 ~Ci/ml) were added~ The cell~ were incuba~ed at 37~C

for 60 minutes. A 10~ aqueous trichloroacetic acid solution was added to the incubation medium to stop the reaction and simultaneously precipitate acid-insoluble matter, The acid-insoluble matter was washed three times with a 5 to 10~
aqueous solution ~f trichl~roacetic acid and then dissolved in formic acid. The radi~actlvity o~ the acid-insoluble matter was ~easured. From the Iati~ of incorporated radiation to hat of a control group, the concentrations which inhibited radiation by 10~, 50~ and 90~ respectively were measured.
The results are shown in ~able 1 below.

TABLE l ActivitY of the compounds of the invention to inhibit the growth, D~A synthesis and RNA synthesis of cultivated leukemic cells L1210 of mice _ . . . . _ _ ~
IC50t~9/ml) IC10(~9/mg) ICgo(~g/mg) COMPOUND 1 day 2 days DNA RNA DNA RNA DNA RNA
later later _ _ _ _ _ Daunomycin 0O049 0.036 0.3 0.1 ¦ 0.072 0.03 1.8 1.8 Adriamycin 0.05 0.03 1.6 0.6 ¦0.19 0.12 4.5 6.5 4-Demetho~y- 0.01 0.005 0.0 0.1 ¦0.007 0.017 1.2 1.4 daunomycin ll-deoxy- 0.05 ____ __ __ __ __ __ daunomycin 4-Demethoxy- 0502 0.006 1.6 1.2 0.12 0.15 10 6.0 adriamycin ll-Deoxy- 0.005 ____ __ __ __ __ __ adriamycin ___ . _ ._ _ __ 4-Demethoxy-ll-deoxy-daunomycin (75,9S,l'~isomer 0.~9 0.01 0.22 0.34 0.02 0,09 2.5 2.5 (7S~9S,l'B)isomer 1.4 n.51 3.1 5.4 0.46 0.9 15 14 (7R,~R,l'~isomer >2.5 >2.5 5.4 9~0 0.8 0.9 13 >10 (7R,9R,l'g)is~mer 2.0 1.25 4.5 5.0 0.6 0.8 15 13 4-Demethoxy-ll-devxy-~driamycin ~75,95,1'~)isomer 0.07 0.03 0~64 0.~ 0.07 0.08 5.6 4.6 (7R,9R,l'~)isomcr 72.5 72.5 ~10 >10 7.4 7.4 >10 >10 ~ . . . ~

9~

B. Antitumor activi.y on CDFI mouse leukemia induced b mouse L1210 leukemia cells Y

L1210 leukemia cells of mice were intraperitoneally transplanted in an amount of lxlU5 cells~mouse in CDFI mice.
Starting 24 hours after the transplantation 9 each of the test compounds was intraperitoneally administered to the mice for 10 consecu~ive days. The survival ra~e ~T/C, %) was calculated in comparison with a control group (to which physioloqical saline was administered). The results are shown in Table 2 below.

Antitumor Activity ~T/C, %) _-Dosage ~g~k~day,mouse)l ~ ¦ Surv: val ratl (T/C, %) Compound ~~~-~_ 5 2 5 1 25 O.6 O.3 0.15 I ~a, O _ Daunomycin ¦throug} l138 191 145 132 118 Adriamycin ¦ 189* 351 272 239 1~7 130 4-Demethoxy~
deoxydaunomycin t7S,gS,1') 1 -- 1~0 135 115 115 103 4-Demethoxy~
deoxyadriamycin (7S,9Srl'~) ~ -- 177* 308 184 162 135 *Toxicity As can be seen from the above experimental results 9 the compounds of ~or~u~a I pro~ide~ ~y the present i~vention, expecially those in which Rl i6 hydrogen or hydroxyl, exhibit excellent antitumor activity on L1210 leukemic cells and experimental animal tumors. The antitumor activity of the 4-demethoxy-ll-deoxyadriamycin (7S,9S~ ) stereoisomer is par-ticularly noteworthy.
The compounds of formula X provided by the present ~9~9~

invention are also characterized by thei antimicrobial activity. Ta~le 3 summarizes the minimum inhibitory con-centrations (MIC) of representa~ive compounds ~f the present inventi~n against various microorganisms on a nutrient agar medium.
~ABLE 3 _ __ MIC on various ~acteria - - - - - - -- _nr ~ . ._ . _ M, :.C. (~g/ml) ~ . .

~ac-teria ompound A ComPound B Com~ound C Com~ound D
. . ...... ~ .. _ .
Staphylococcus aureus ~DA 209 25 >100 100 >100 . . .__ .. ._ Staphylococcus aureus smith 12.5 _>100 12.5 >100 Bacillus subtilis PCI 219 3.12100 12 5 >100 . . ,__ .__ _ __ Bacillus subtilis NRRB- 558 6.2550 12.5 >100 _ ..... _ ._ . . ...
Bacillus cereus ATCC 10702 12.5 50 __ 25 >100 ~acillus megather~
ium APF ~ _6.2550 _ 12.5 _>100 Sarcina lutea PCI 1001 6.2550 100 >100 . _ . ''----- '''---I
Microccus flavus FDA 16 _ ~.25>100 50 >100 _ ¦

Microccus lysodeik-ticus IFQ 3333 12.5 >100 50 >100 . __ . _ _. . .___ .

Corynebacterium bovis 1810 6.25100 100 >100 ._ ......... . _ . _ _ .
Klebsiella pneu-monia PCI 602100 >100 >100 >100 . .__~ . _ .
Escherichia coli NI~J >1~0 >1~0 ~lD0 >100 . . . _ _ _ . . _ __ Salmonella ~yphi T - 63 >100 >100 >100 >100 .__ _ .. _ . _ _ .
Shigella flexn~ri.
46 JS 11811 lO0 >100 >100 >100 --_ .. .. _ _ __ _.... _ _ Pseudomonas aeruqinosa A3 >100 ~100 >100 >100 -~--~ _ ~_ .. _ _ .... .. _. _ ._ Candida albicans 3147 100 _ _ >10~ >100 100 _ __ ~_ - 19 -9~9~
TABLE 3 - continued __ Mycobacterium smegma _ tis ATCC 607 12.5 100 12.5 100 _ ompound A: 4-Demethoxy 11-deoxydaunomycin, ~7S,9S,l') isomer ompound B: 4-Demethoxy-ll-deoxydaunomycin, (7P~,9R,l') isomer ompound C: 4-Demethoxy-ll deoxyadriamycin, t7S,9S,l'~) isomer ompound D: 4-Demethoxy~ deoxyadriamycin, (7R,9R,l'~) isomer Therapeutic Use The compounds of formula I and their nontoxic acid addition salts possess activity as antimicrobial agents, use-ful in both human and veterinary medicine, and also marked inhibitory action against malignant mammalian tumors, includ-ing both solid ~nd ascitic types, According to one aspect of the invention, a method is provided for therapeutically treating a mammalian host affected by a microbial infection (particularly a gram-positive bacterial infection) or by a malignant tumor (i.e. a solid-or ascitic-type tumor such as L1210 leukemia) which comprises administering to said host an effective antimicrobial or tumor-inhibiting dose of a compound of formula I, or a nontoxic acid addition salt thereof, According to another aspect of the invention, a pharmaceutical composition is pro~ided which comprises a therapeutically effecti~e antimicrobial or tumor-inhibiting amount of a c~mpound Df formula ~, or a nontoxic acid addition salt thereof, in combination with a compatible pharmaceutical carrier or diluent, Preferably, such compositions ~re made up in a form appropriate for paren~eral administration, Preparations f~or parenteral administration include sterile aqueous or non-aqueous solutions, suspensions or emulsions. They may also be manufactured in the form of sterile solid compositions which can be dissolved in sterile water, physiol~gical sali~e or some ~ther 5terile injectable medium immediately before use.
It will be appreciated khat the actual preferred dosage amounts used will vary according to the particular compound belng used, the particular composition formulated, the mode of application and ~he particular situs, host and disease being treated In general the compounds are admin-istered intraperitoneally, subcutaneously, intravascularly (intravenously or in~raarterially) or topically to non-human mammals and intravascularly or topically to humans. Many factors that modify the action ~f a drug will be taken into account by those skilled in the art, for example, age, body weight, sex! diet, time of administration, route of adminis-tration, rate of excretion, con~ition of the patient, drug combinations, reaction sensitivities and severity of the disease. Administration may be carried out continuously or periodically within the maximum tolerated dose, Optimal dosages and application rates for a glven set of conditions can ~ ascertained by those skilled in the art using con-ventional dosage ~etermination tests based on the above guidelines. Typically, the dosage will be about O, 2 to 5 mg/kg of body weight.
For use as an antimicrobial ag~nt, the compounds are in general administered so that the roncentration of active ingredien~ is greater than the minimum inhibitory concentration ~r the par~icular ~gaDis~ being tr~ated The number o~ administr~ti~s, d~s~g~ ~rm, etc. may be easily determi~ed b~ t~e skilled per~ using conventional dosage determination tests.
The following examples are pro~ided for illustra-tive purposes only and are not intended to limit the scope of the present inven~ion, In ~he ~xamples~ the ratio of solvent .. .. .. .....

in amixture is indicated in the ration of volume to volume.

Prodiction of 4-demethoxy-11-deoxydaunomycino e Step a 2-Ethinyl-2-hydroxy-5-mehtoxy-1,32,3,4-tet4ahydroaphtalene:- (2) While acetylenbe was blown into 80 ml of anhydrous tetrahydrofuran, 36m; of a 2M ehter solution of ethylmagnesium bromide was added dropwise.
To the resulting solution was added 2.88 g of 5-mehoxy-2-terralone. After the reaction, thd reaction mixture was poured ihnto 500 ml of a saturated aqueous solution of ammonium chlorice, and the mixture wss extracted with 200 ml of carbon tetrachloride three times. The extracts were washed with water, dried over sodium sulfate, and concentrated to dryness. The resulting bronw oily product was chromatographed on a colunm of silica gel using benzene/ethyl acetate (25/1) as an eluent to gie 1.6 g of the desired product as a brown oil.
NMR = 60MHZ, CDCl3 1.9 - 2.3 (m) -CH2_at 3-position 2.4 0 (s) -C= CH at 2-position 2.4 2 (s) -OH at 2-position 2.7 - 3.1 (m) -Ch- at 4-position 3.05 - 3.2 (m) -CH- at 1-position 3.8 2 (s) -OCH3 at 5-position 6.6 - 7.35 (m) H at 6-, 7-and 8-positions Step B
2-Acetyl-2-hydroxy-5-mehoxy-1,2,3,4-etrahydronaphthalene:- (3) tHE compound of formula (2) was dissolved in 10 ml of carabon tertrachloride, and 10 ml of 1.5N sulfuric acid and 200 mg of mercuric oxide were added. They were reacted at room tempreature for 27 hours. Water (40 ml) was added, and he reaction mixture was extracted with 50 ml of cargon tetra-chloride four times. Thje extracts were washed with water, dried over sodiium sulfate, and concentrated on a colume of resulting oily product was chromatographed on a column of silica gel using benxene/ethyl acetate (20/1) as na eluent to give 840 nmg of the desired product as colorless needles.
Melting point: 63 - 63.5 C
NMR: 60 MHz , CDCL3 1.8 - 2.1 (M) -CH2- AT 3-position 2.28 (s) COCH3 at 2-position 2.5 - 3.5 (m) -CH2- AT 1--AND 4-positions 3.45 (s) OH at 2-position 3.83 (s) OCH3 at 5-position 6.6- 7.3 (m) H at 6-,7- and 8-positions Step C
4-Demethoxy-7. 11-dideoxydanomycinone:- -969~4 230 mg of the compound (3) obtained in step B, 230 mg of phthalic anhydride, 46D mg ~f sodium chloride and 2.3 g of alumin~m chl~ride were well mixed, and melted by heating the~ ~ ~80C. In 1 to 2 minutes, the mixture became homogeneous, but it was reacted further for ~ minutes, The reaction mixture was treated with a saturated aqueous solution of oxalic acid, and extracted with 30 ml of chloroform five times. The extracts were combined, washed with water, dried over sodium sulfate, and concentrated to dryness. The product was chromatographed on a column of silica gel using benzene~ethyl acetate (20/1) as an eluent to give 110 mg of the desired product as a yellow solid.
Melting point: 208 - 214C (decomp.) NMR~: 60 MHz , DMSO
107 2.2 (m) -CH2- at 8-position 2.30 (s) -COCH3 at 9-position 2.6 3.1 (m) -CH2- at 7-and 10-positions

5.60 (s) -OH at 9-position 7.38 (s) -H at 11 position 7.8 8.4 (m) -H at 1-,2-,3- and 4 positions 12.80 (s) -OH at 6-position Step D
Ketal derivative of the compound (4):-O
O \C''C~3 OH

HO
(~-a) 100 mg of the compound (4) was dissolved in 15 ml of benzene, and 0.3 m~ of ethylene glycol and a catalytic amount of p-toluenesulfonic acid were added. The mixture was reacted for 3 hours under reflux. The reaction mixture was then poured into a 0.02N aqueous solution of sodium hydrogen carbonate and extracted with 40 ml of ethyl acetate three times. The extracts were dried over sodium sulfate, and concentrated to dryness. The above reaction proceeded quantitatively.

Step E
4-Demethoxy-ll-deoxydaunomycinone:-O 1~

O HO OH

The ketal compound (8-a) obtained in step D was dissolved in chloroform, and 12,6 ml of a 0.8% (W~V) bromine in carbon tetrachloride solution, 21 ml of water and 120 mg of azobisisobutyronitrile were added. The reaction mixture was reacted at room temperature for 3.5 hours. After the re-action, 4 ml of the above ~romine solution was further added, and the reaction was carried out for 2 hours. After the re action, bromine, carbon tetrachloride and chloroform were distilled off from the reaction mixture, The residue was dissolved in 120 ml of acetone, and 21 ml of 8N hydrochloric acid was added. The reaction was carried out at room temper ature for lB hours IdeProtection of the ketal, and the epimer-ization of (7S,9R; 7R,9S~ to ~,9S; 7R,9R~]. After the reaction, acetone was distilled off from the reaction mixture, and the r~sidue was extracted with 25 ml of _hloroform twice.
The extracts were washed with water, dried over sodium ~ulfate, and concentrated to dryness.

The product was treated with a crosslinked dextran qel (for example, Sephadex*LH-20, a ~radename for a product of Pharmacia F-ne Chemicals~ using acetone, and then the acetone was distilled o~ t~ gi~e ~he desired product I a mixture of a racemic mixture (7S,9R; 7R,9S) and a racemic mixture (7S,9S; 7R,9R)~. The above stereoisomeric mixture was chromatographed on a column of silica gel using benzene/ethyl acetate (16/1), benzene/ethyl acetate (12/1), benzene/ethyl acetate ~8/1) and then benzene/ethyl acetate (4/1) to give 41 mg of a stereoisomer (7S,9S; 7R,9R) of the above compound as a yellow solid and 21 mg of a stereoisomer (7S,9R; 7R,9S) of the above compound as a yellow solid-Step EEpimerization of 4-demethoxy-11-deoxydaunomycinone (7SI9R;
7R,9S) to its isomer (7S,9S; 7R,9R):-24 mg of the above 17S,9R; 7R,9S) isomer was dissolvedin 15 ml of acetone, and 1.5 ml of 60~ perchloric acid was added.
~he reaction was carried out at room temperature for 3 hours~
The reaction mixture was worked up and purified in the same way as in step ~ to give 12 mg of 4-demethoxy~ deoxydauno-mycinone (75,9S; 7R,9R).
Melting point: 199 - 207~C (decomp.
NMR~: 100 MHz CDC13 2.0 - 2.5 (m) -CH2- at 8-position 2.Q4 (s) -C-CH3 at 9-position 3~14 (A~) -CH2- at 10-position 3.6~ -~H at 7-position 4.60 (s) -OH at ~-position 5.36 ~m~ -H at 7~position 7.60 (s) -~ at ll-position 7~7 7.9 (m~ -H at 2-and 3-positions 8.2-8.4 (m) -H at l-and 4-positions 13.23 (s) -OH at 6-position * trade mark.

9~.9~L~

.

.
Production of 14-O-acetyl-4-demethoXy-ll~
~lr~ D~
O O

HO OH

Step A

_4-O-Acetyl-4-demethoxy-7,11-dideoxyadriamycinone:-O O

~' ~ ~ H

HO (6-a) 110 mg of the compound (4) obtained in step C of Example 1 was dissolved in 12 ml of tetrahydrofuran, and 200 mg of N-bromosuccinimide (NBS) was added four times at l-hour intervals. The reaction was carried out at room temperat~re for a total period of 6 hours. The reaction mixture was poured into 200 ml of water, and extracted with 100 ml of carbon tetrachloride two times. The extracts were dried over sodium sulfate, and concentrated to æryness under reduced pressure.
The product, without isolation and purification, was dissolved in 40 ml of acetone, ana 200 mg of potassium acetate was added.
The reaction was carried out at room temperature for 8 hou~sO
After the reaction, the reaction mixture was poured into 50 ml of water, extracted with 25 ml o~ chloroform four times, dried over sodium sulfate, and then concentrated to dryness.
The prod~ct was chromatographed on a column of silica gel using benzene~ethyl acetate (40/1), ~enzene/ethyl acetate (30/1), benzene/ethyl acetate (20~ and then benzene/
ethyl acetate (15/1) as an eluent to give 83 mg of the title compound.

i9~ ~

Melting point: 206 - 209~C (Dec ~ ) NMR~: 100 MHz CDC13 2.0 2.3 (m) -CH2- at 8-position 2.20 ~s) O~ CH

2.79 (s) -OH at 9-position 2 8 3.5 (m) -CH2 at 7-and 10-positions 5.11 (s) ~2 3 7.60 (s) ~ at ll-position 7.75 7.9 (m) -H at 2-and 3-positions 8.2 8.~ (m) -H at l-and 4-positions 13.03 (s) -OH at 6-position Ste~ B

14-O-Acetyl-~-demethoxy-ll-deoxyadriamycinone:-_ 51 mg of the compound (6-a) was dissolved in 14 ml of chloroform, and 12 ml of water and 16 mg of azobisisobutyro-nitrile were added. To the solution was added 0.5 ml of a 0.8%
(W/V) bromine in caxbon tetrachloride solution in seven portions at l-hour intervals at room temperature, and the reaction was carried out for 9 hours. After the reaction, the unreacted bromine, carbon tetrachloride and chloroform were distilled of~. The residue was dissolved in 40 ml of ethyl acetate, and 10 ml of a 0.lN aqueous solution of sodium hydrogen carbvnate was addedO The mixture was stirred at room temperature for a day and night. ~ter ~he ~eaction~ the reaction mixture was sub-jected to a separating procedure. The ethyl acetate layer was ~ashed with water, dried over sodium sulfate, and con-centrated to dryness. The dried product was chromatographed in the same way as in step A to give 34 mg o~ a racemic mixture (7S,9S; 7R,9R) of the title compound~
Melting po~nt: ~98 - 205~C (decomp 9~

NMR~: 100 MH~ CDC13 2O0 2.6 (m) -CH2 - at 8-position 2.19 (s) O~-CH3 3 . 28 -OH at 7-position 3.22 (AB~ -C~2 ~ at 10-position 4.69 (s) -OH at 9-position 5.25 (AB~ -~-CH2-O~C~3 5.42 (m) -H at 7-position 7.68 (s) -H at ll-position 7.75 7.9 (m) -H at 2-and 3-positions 8.25 .8.4 (m) -H at l-and 4-positions 13.40 (s) -OH at 6-position IR (cm ~) 3450, 2950, 1750, 1730, 1670, 1630, 1590, 1480, 1420, 1385, 1360, 1330, 1285, 1270, 1245, 1215, 1160, 1110, 1100, 1065, 1060, 1040, 1030, 1010, 980, 960, 930, 910, 870, 840, 830, 820, 795, 770, 745, 715.

Production of 4-demethoxy-11-deoxydaunomycin_ ~1 C-C~

r ~H2 HO

Step A
4-O-p-nitrobenzoyl-1,2,3,6-tetradeox~-3-trifluoroacetamide-L-lyxo-hex-l-enopyranose:-2 ~ ~
HCOCF3(1) 10~ mg of N-trifluoroacetyldaunosamine was dissolved in anhydrous pyridine, and 300 mg of p-nitrobenzoyl chloride was added. With ice cooling (about 8DC), the reaction was carried out for 22 hours. After the reaction, the reaction mixture was poured into 100 ml of ice water to destroy the excess reagent. The reaction mixture was then extracted with 30 ml of chloroform three times. In order to remove pyridine from the extracts, the extracts were subjected to a separating procedure using 50 ml of lN hydrochloric acid. The chloroform layer was washed with water, dried over sodium sulfate, and concentrated.
~2) The concentrate (crude 1,4-bis-O-p-nitrobenzoyl-N-trifluoroacetyldaunosamine) was dissolved in 10 ml of acetone, and 5 ml of 4N hydrochloric acid was added. The reaction was carried out at room temperature for 10 hours to partially hydrolyze the p-nitrobenzoyl groupO After the reaction, the acetone was remo~ed by concentration, and 15 ml of water was added. The mixture was extracted with 10 ml of chloroform three times. The extracts were subjected to a separating procedure using 10 ml o a 0.lN aqueous solution of sodium hydrogen carbonate. The chloroform layer was washed with water, dried over sod-um sulfate, and concentrated.
(3) The concentrate ~crude 4-O-p-nitrobenzoyl-N-tri~luoro-acetyldaunosamine) ~7as dissolved ïn lS ml o~ anhydrous pyridine, and 600 mg of p-toluenesulfonyl chloride was a~ded.
The reaction was carried ou~ at 80~C for 18 hours. The light brown reaction mixture was poured into 100 ml of ice water to destroy the excess reagent. The reaction mixture was then 9~

extracted with 30 ml of benzene four tirnes. The extracts were washed with water, dried over sodium sulfate, and concentrated. The oily produc~ was chromatograph~d on a column of silica gel using benzene/ethyl acetate (25/1) as an eluent to give 118 mg of the title compound.
Melting point: 144 - 148C
[~]D : -100~C (c: 0.5, acetone) NMR~: 100 MHz CDC13 1.32 (d)-CH3 at 6-position 4.36 (broad q)H at 5-position 4.62 (td)H at 2-position 5.05 (m)H at 3-position 5.66 (broad d)H at 4-position

6.15 (broad d)-NHCOCF3 6.65 (dd) H at l-position 8.15 8~4 (m) H ~ H

H H
Step B
4-Demethoxy-4'-O-p nitrob_ zoyl-ll-deoxy-3'-N-trifluoro-acetyldaunomycin-- O

O HO

~3 ~COCF3 (I-a dco~}~2 20 mg of the 4-demethoxy-11-deoxydaunomycinone (7S,9S; 7RI9R) obtained by the method of Example l and 80 mg of glyc~l o~tained in step A above were dissolved in 2 ml of benzene. A catalytic amount of p~toluenesulfonic acid was added, and the reaction carried out at room temperature for 93 hours. After the reaction, the reaction mixture was poured into 30 ml of a 0.01N aqueous solution of s~dium hydrogen carbonate, and extracted with 15 ml of benzene three times.
The extracts were washed with water, dried o~er sodium sulfate, and conce~trated to dryness. The product was chromatographed on a silica gel column 1lsing benzene/ethyl acetate (4/1) as an eluent, further chrvmatographed on a column of a crosslinked dextran gel ~e.g. Sephadex LH-20) using chloro~orm/acetone ~1/2) as an eluent; and further chromatographed on a column of silica gel using benzene/ethyl acetate (4/1) as an eluent to isolate the individual stereoisomers of the title compound. The isolated crude products were again treated individually with Sephadex LH-20 to give 7 mg of a (7S,9S,l'a) isomer (I-a~
of the title compound, 3 mg of a (7S,9S,l'~) iscmer (I-al-2) of the title compound, 10 mg of a (7R,3R,l'~) isomer (I-al-3) of the title compound, and 3 mg of a (7R/9R,l'~) isomer (I-al-4) of the title compound.

( I-al-l) Melting point: 153 - 156C
[~]D4: -125C (c: 0.2, acetone) NMR~: 100 MHz CDC13 1.27 ~s) -CH3 at 6'-position 2.0 2.5 ~m) -CH2 at 8-and 2'-positions 2.43 (s) -COCH3 at 9-position 3.21 (AB) -CH2 at 10 position 4.25 (s) -OH at 9-position 4.3~4 7 (m) ~~ at 3' position 4O4g ~r~a~ H at 5i-position 5,34 (m) -H at 7 -position S.51 (m) -H at 4'-position 5.69 (~I) H at l'-position 69~L~

6.40 (broad d) NHCOCF3

7 . 64 (s) -H at ll-position 7.7 7.9 (m) -H at 2-and 3-positions

8.2 8.45 (m) ~ -H at l-and 4-positions H
NO

H
13.33 ~s) -OH at 6-~osition (I-al-2) Melting point: 142 - 145DC
[~]D : +87.5 (c: 0.2, acetone) NMR~: 100 MHz CDC13 i.o7 (d) -CH3 at 6 ' -position 1.7-2.6 (m) -CH2 at 8-and 2'-positions 2.39 (s) -COCH3 at 9-position 3.27 (broad s) -CH2 at 10-position 3.84 (broad q) -H at 5'-position 4.2~4.6 (m) -H at 3'-position 4.74 (broad s) -OH at 9-position 5.1.5 (dd) -H at l'-position -: . 5.32 ~m) -H at 4'-position 5.66 (m) -~ at 7-position 6.54 (broad d) NHCOCF3 7.61 (s) -H at ll-position 7.75 7,9 (m) -H at 2-and 3-positions 8.1 8.4 (m) ~ -H at l-and 4-positions ~ H ~ H
IC~02 ~ H

13.36 ~s3 OH at 6-position (I al 3) Melting point: 148 - 152C
1~25: -225C (c: 0.2, acetone) NMR~: 100 MHz CDC13 1.26 (d) -CH3 at 6'-position 1. 6 2 . 7 (m~ - CH2 at 8-and 2'-positions 2 . 39 ~s~ -COCH3 at 9-position 3.24(broad 5) -CH2 at 10-position 4.52 ~s) -OH at 9-position 4O45~4~8 (m~ -H at 3'-position 4. 73 (broad q) -H at 5 '-position 5.44 (m) -H at 4'-position 5.5~5.57 (m) -H at 7-and l'-positions 6.62 (broad d) -NHCOCF3 7.61 (s) -H at ll-position 7.75~7.95 (m) -H at 2-and 3-positions 8.2~8.4 (m) ~ -H at l-and 4-positions / I~E~

~ n H

13.43 (s) -OH at 6-position ( I-al-4 ) Melting pointo 190 - 195~C
[~]25: -137.5 (c~ 0.2, acetone) NMR~: 10~ M~z CD~13 1.32 (d) -CH3 at 6'-position 1. 6 ~20 85 (m) -CH~ at 8-and 2 ' -posi~ion~
3.17 (AB) -CH2 ak 10-position 4 . ()1 (broad q) -~ at 5 ' ~position 4 . 25 ~4 . 6 (m) -H at 3 ' position 4 . 55 (s~ -OH a~ 9~positic~rl 5.20 (dd) -H at l'-position 5.3 5.45 (m) -~ at 7-and 4-positio~s 6.59 (broad d) -NHCOCF3 7.57 (s) -H at ll-position 7.7 7.9 (m) -H at 2-and 3-positions 8.15 8.4 (m) ~ -H at 1-and 4-~csiti~ns ~ H ~ H
¦-CQ< O ~N02 >~
B H
13.25 (m) -OH at 6-position Step C

4-Demethoxy-ll-deoxydaunomycin -O O

O HO ~o 3 ~ ~
~ NH3. (I-l) 12 ~g of N-trifluoroacetyl-4'-O-p-nitrobenzoyl-daunomycin (?S,9S/l'a~ (I-a~ was dissolved in methanol, and 1 ml of a 10~ aqueous solution of potassium carbonate was added. Under ice cool~n~ ~about ~C), the reaction was carried out for 12 hours. After the re~ction, 15 ml of water was added, and the r~ac~ion mixtu~ ~as extracted with 10 ml of chloro-form foux t~mes~ The ch?oro~orm layer was treated with 5 ml of 0.5% aqueous acetic acid four times. The acetic acid layer was neutralized with a lN aqueous solution of sodium hydrogen carbonate, and a~ain extracted with chloroform. The extract was dried over sodium sulfatet and concentrated to dryness.
The dried solid was dissolved in dichloromet~lane/t-butanol (1/20) and lyophilized to gîve 7.5 mg of a l75,9S,l'~ isomer (I-la) of the title compound as a y~llow powder~

~69~

In the same way as a~ove, 4.2 mg of a ~7s~9s~lle) isomer ~I-lb) o~ the title compound was prepared from 7.9 mg of ~I-al-2); 6.5 mg of a ~7R,9R,l'a) isomer (I-lc) of the title compound, from 11 mg of (I-al-3); and 3.6 mg of a (7R,9R,l'B) isomer (I-ld) of the title compound, from 6.3 mg of (I-al-4).
(I-la) Melting point: 202 - 212DC (D~x~) l~]24: ~50~ (co 0.1, methanol) FDMS. MH 482 NMR~: 100 MHz CDC13 1.28 (d) CH3 at 6'-position 1.6 2.6 (m) -CH2 at 8-and 2'-positions 2.39 (s) -COCH3 at 9-position 3.0 3.2 (m) -H at 3'-position 3.24 ~AB) -CH2 at 10-position 3,44 (m) -H at 4'-position 4.28 ~broad q) -H at 5'-position 5.36 (m) -H at 7-position 5,55 (m) ~~ a~ l'-position 7~64 (s) -H at 11-position 7O75 7,95 (m) -H at 2-and 3-positions 8.25 8.45 (m) -H at l-and 4- positions IR ~cm 3450j 2310, 1705, 1665, 1~30, 1590, 1515, 1480, 1455, 1920 13~5, 1355~ 1325, 1~00, 1275, 1250, 1200, 1115, 1005, 980, ~0; 87~, 8~S, 79~, 770~ 715, 6.9~9~

(I-lb) ._ Melting point: 160 - 172C (decomp.) [~D o ~500 (c: 0.1, methanol) Fr)MS: Ml~ 4 ~ 2 NMR~: lOD M~z CDCl 3 1. 25 (d) -CH3 at 6'-position 1. 55 2 . 7 ~m) -CH2 at .8-and 2 ' -positions 2 . 4 9 ( s ) -COCH3 at 9 -pos ition 2.95 3.2 (m) H at 3'-position 3.3 3.45 -CH2- at 10-position -H at 4'-position 3.60 (broad q) -H at 5'-position 5.00 (dd) -H at l'-position 5 . 6 8 Im) -H at 7-position 7 . 61 (s) -H at ll~position 7 . 7 5 7.9 5 ~m) ~H at 2-and 3-positions 8, 2 8 . 4 (m) -H at l-and 4-positions I F~ ( C~T ) ~400, 29DG, 1705, 1670, 1630, 1590, 1480, 1420, 1385, 1360, 1330, 1300, 1275, 1255, 120~, 1165, 1115, 1060, 980, 920, 870, 830, 775, 720.

( I - 1 c ) Melting point: 180 ~ 195 DC (decomp.1 ~D : -150D (C: 0.1~ m~thanol) FD~S: ~ 4P,2 NMR~: lOû MHz CDC~3 35 (d~ -CH3 a~ 6 ' -position 1~6-20~ (m) -CH2 at 8-and 2'-positions 2.40 (s) -COCH3 at 9-position 2.9-3.2 (m) -H at 3'-position 6.9~

-3.21 (AB) -CH2 at 10-position 3. 4B (m) -H at 4'-position 4.11 (broad g) -H at 5'-position 5.32 (m) -H at 7-position 5.50 (m) -H at l'-position 7.65 (s~ -H at ll-position 7,75 7,g (m) -H at 2-and 3-positions 8.25 8.45 (m) -H at l-and 4- positions IR (cm 3450, 2900, 1710, 1670, 1630, 1590, 1480, 1420, 1385, 1360, 1330, 1300, 1270, 1250, 1200, 1160, 1120, 1080, 1010, 980, 930, 875, 825, 790, 770, 715.

(I-ld) Melting point: 185 195C (decomp.) [a]D : ~400 (c: 0.1, methanol FDMS: MH 482 NMR~: 100 ~Hz CDC13 1.39 (d) -CH3 at 6'-position 1.35 2.85 (m) -CH2 at 8-and 2'-positions 2.42 (s) -COCX3 at 9-position 2.9 3.2 (m) ~H at 3l-position 3.15 (AB) -CH2 at 10-position 3,33 (m) -H at 4'-position 3.66 ~broad q1 -H at 5'-position 5.00 (dd) ~~ a l'-position 5.33 ~ at 7~position 7.~4 ~s~ -~ ~t ll-positi~D
7.7 7.~ Im) -H at 2-and 3-positions 8~2-8.4 (m) -H at l-and 4-p~sitions IR (cm 3450, 290~, 1710, 1670, 1630, 1590, 1~0, 1420, 139C, 136 g6~
- ~8 -1330, 1300, 128~, 1255, 1210, 1170, 1130, 1110, 1065, 1025, 985, 920, 870, 830, 790, 7B0, 720.

Production of 4-demethoxy~ deoxyadriamycin ~I CCH2~H
~ H

o HO ~d ~ //
3 ,/ r NH2 (I-2) OH

Step A
14-O-acetyl-4-demethoxy-4~-o-p~nitrobenzoyl-ll-deoxv~-3~-N
trifl~oroacetyladriamycin:- _ O
o ~I Ç-CH3 r 0 ~30 o NHCOCF3 (I_~
i-C0 ~ 2 32 mg of the 14-~-acetyl--4-demethoxy-11-deoxy-adriamycinor~e ~7s,9s; 7R,g~ obt~ined ~y the rnethod of Example 2 and 12~ mg o the glycal obtained in step A of Example 3 were dissolved in 1~ m~ of aichloromethane. A cataly~ic amount of p-toluenesul~onie acid was added, and ~he reaction was carried out at room temperature for 75 hours. After the re-actiori, the reaction mixture was poured into 10 ml o~ a 0.01N
aqueous solution of sodium hydrogen car~onate, and ~as g~

extracted with 10 ml of dichlor~methane two times, The extracts were washed with water, dried over sodium sulfate, and concentrated to dryness, The product was worked up in the same way as in the purifying procedure in step B of Example 3 to give 10 mg of a t7S,9S, 1'~ isomer (I-a2-1) of the title compound and 10.7 mg of a (7R,9R, 1'~) isomer (I-a2-2) of the title compound, (I-a~-l) Melting point: 154 - 157C
[~]25 125 (c: 0.2, acetone3 NMR~: 100 MHz CDC13 1.31 (d) -CH3 at 6'-position 1.9 2.7 (m) -CH2 at 8-and 2'-positions 2.22 (s) OCOCM3 3.31 (AB) -CH2 at 10-position 4.41 (s) -OH at 9-position 4,3 4,7 (m) -H at 3'-position 4.45 (broad q) -H at 5~-position 5.24 (AB) CCH2 3 5,40 (m) -H at 7-position 5 50 (m) -H at 4 '-position 5,70 (m) -H ~t ll-position 6,30 (broad d) -NH-COCF3 7.69 ~s) -H at ll-position 7.75~7.95 lm) ~ at 2-and 3 positions 2-8.~5 lm) ~ H at l-and 4-positions ~ H H

//CO ~NO 2 13.38 ~s) -OH at 6-position 36.~

( I-a2-2 ) Melting point: 155- 160~C

lc~25: -22~ (c: 0.2, acetone) NMR~: 100 MHz CDC13 1.26 (d3 -CH3 at 6'-position 1.8 2.8 (m~ -CH~ at ~-and 2'-positions 2.20 (s) OCOCH3 3.31 (broad s) -CH2 at 10-position 4.4 4.85 (m) -H at 3'-position 4.72 ~broad q) -H at 5'-position 4.62 (s) -OH at 9-position 5.21 (AB) CO ~H2 OCOCH3 5.41 (m) -~ at 4'-position 5.5 5065 (m) -H at 7-and l'-positions 6.52 (broad d) NH-COCF3 7.63 (s) -H at ll-position 7.75 7.95 (m) -H at 2-and 3-positions 8.2 8.4 (m) ~ -H at l-and 4-p~sitions H
--CO~No2 . ~
13.46 (s) -OH at 6-position Step B
4-Demethoxy~ deoxyadriamycin.-(1) 1?.3 xng o~ the compound ~I-a2-1) o~tained in step A was dissolved in 6 ml ~f methanol, and 0.06 ml of a 10~ aqueous solution oE potassium carbonate was addedO The reaction was carried out at 0C for 3 hours. Ater the reaction, 30 ml of water was addedt and extracted with 10 ml of chlor~Eorm three 69~

s-~lEate, and concentrated to dryness. The dried product was crude N-trifluoroacetyl-4-demethoxy-11-deoxyadri~mycin (7S,9S, 1 ' - a ) .
t2) The dried crude product was dissolved in 3 ml of dichloro-methane, and 1.2 ml of triethoxyme~hane and a catalytic amount of p-toluenesulfonic acid were added. The reaction was carried out at room temperature for 2.5 hours. After the re-action, lS ml of a O,lN aqueous solution of sodium hydrogen carbonate was added to neutralize the reaction mixture, followed by extraction wikh dichloromethane. The extract was dried over sodium sulfate, and concentrated to dryness.
(3) The resulting dried product was dissolved in 4 ml of methanol, and 1 ml of a 10% aqueous solution of potassium carhonate was added. With ice cooling (about 8C), the re-action was carried out for 16 houxs. After the reaction, 15 ml of water WhS added, and the reaction mixture was extracted with chloroform. Th~ extract was treated with a 1% aqueous solution of acetic acid (when the orthoformate was eliminated).
The acetic acid layer was adsorbed onto an adsorbent resin (e.g. Amberlite XAD-2, a tradename for a product of Rohm h Haas Co.), and eluted stepwise with acetone/O.OOOlN hydrochloric acid (20/80, acetone/O.OOOlN hydrochloric acid (30/70~, and then acetone/O,OOOlN hydrochloric acid (40/60). After dis-tilling off the acetone, the eluates were lyophilized to give 3.0 mg of the hydrochloride of a (75,9S/l'~ isomer (I-2-1) o 4-demethoxy-11-deoxyadriamycin as a yellow powder.

9.8 mg of the compound (I-a2 2) obtained in step A
was similarly treated to give 2~5 mg of the hydrochloride of a ~7R,9R,1'~) isomer (I-2-2~ of 4-demethoxy-11-deoxyadriamycin.

H drochloride of Melting point: 158 ~ 170C ~decomp.
l~24 ~75 (c: 0.1, water) * krade mark.

F~ MH 49~
NMR~: 100 MHz D2O
1.80 (d) -CH3 at 6'-position 2.4 2,9 (m) -CH2 at 8-and 2'-positions 3,45 (broad s) -CH2 at 10-position 4.1 4.4 (m) -H at 3'-position 4.33 (m) -H at 4'-position 4.66 (broad q) O -H at 5'-position .30 (s) -C-CH2-OH
5.94 (m) -H at l'-position 7.53 (5) -H at ll-position 8.2 8.5 ~m) -H at 1 ,2-,3-and 4-positions IR (cm 3400, 2900, 1720, 1670, 1630, 1590, 1510, 1480, 1420, 1385, 1360, 1330, 1300, 1275, 1250, 1200, 1115, 108~, 1060, 1010, 980, 9~0, 910, 875, 825, 770, 715.

Hydrochloride of ~I-2-2) Melting point: 145- 155C (decomp.) [~]D : 125~ (c: 0.1, water) NMR~: 100 MHz D2O
1.73 (d) -CH3 at 6'-position 2.3 3.2 (m) -CH2 at 8~and 2'-positions 3O50(broad s) -CH2 at 10-position 4~1-4.3(m~ -~ at 3'-position 4.32(m~ -H ht ~ ~-position 4.~~broad ~ H at 5'-position 33 (s~ ~C~CH2-OH
5.47 (m) ~~ at 7-position 5.91 (m) -H at l'-position 7~48 (~ -H at ll~position ~6.~

8.2~8~5 (m) -H at 1-,2-,3-and 4-positions IR ~cm 1) 3450, 2900, 1720, 1670, 1630, 1590, 1480, 1425, 1390, 1365, 1330, 1300, 1280, 1255, 1195, 1110, 107S, 1010, 9~5, 935, 910, 830, 790t 770, 715, EXAMPLE S
Salt Formation Illustrative of the procedures which may be used to prepare acid addition salts, the free base of 4-demethoxy-ll-deoxyadriamycin (7S,9S,l'~) and 4-demethoxy-11-deoxydauno-~ycin (7S,9S,ll a ) may be dissolved in ethyl acetate and about one equivalent of ~Cl added. On lyophilization, the appropriate hydrochloride salt is obtained.
Acid addition salts of ~.he other anthracycline glycoside compounds of formula I may be prepared in a similar manner by using the appropriate organic or inorganic acid and appropriate free base starting material.

Claims (21)

We claim

1. A process for producing an aglycone intermediate of the formula wherein R1 represents hydrogen, hydroxyl or (lower) -alkanoyloxy, which comprises the steps of A) subjecting a compound of the formula wherein Y represents -C-CH2-(lower)alkanoyloxy or in which Z is a ketal residue and W represents a halogen atom, to hydrolysis;

or where the intermediate of the formula is desired; the alternative step of B) reacting a compound of the formula.

with phthalic anhydride of the formula in the presence of ? ????? ?????? ?????????

2. A process for producing an intermediate of the formula which comprises reacting a compound of the formula with phthalic anhydride of the formula in the presence of a Lewis acid and in the absence of a solvent at a temperature of at least 150 C.

3. A process for producing an aglycone intermediate of the formula wherein R1 represents hydrogen, hydroxyl or (lower) -alkanoyloxy, which comprises the steps of A) subjecting a compound of the formula wherein Y represents CH2-(lower)alkanoyloxy or in which Z is a ketal residue and W represents

4. The process according to Claim 3 wherein Z is selected from the group consisting of

5. The process according to Claim 3 or Claim 4 in which the aglycone product is recovered in the form of the racemic mixture (7S, 9S; 7R, 9R).

6. An aglycone intermediate of the formula where R1 represents hydrogen, hydroxyl or (lower) -alkanoyloxy, whenever prepared or produced by the process of claim 3 or by an obvious chemical equivalent thereof.

7. The process according to Claim 3 wherein R1 is hydrogen.

8. The process according to Claim 3 wherein R1 is hydroxyl.

9. The process according to Claim 3 wherein R1 is (lower) - alkanoyloxy.

10. The compound according to Claim 6 wherein R1 is hydrogen, whenever prepared or produced by the process of Claim 7 or an obvious chemical equivalent thereof.

11. The compound according to Claim 6 wherein R1 is hydroxyl, whenever prepared or produced by the process of Claim or an obvious chemical equivalent thereof.

12. The compound according to Claim 6 wherein R1 is (lower)-alkanoyloxy, whenever prepared or produced by the process of Claim 9 or by an obvious chemical equivalent thereof.

13. An intermediate of the formula whenever prepared or produced by the process of claim 2 or by an obvious chemical equivalent thereof.

14. A process for producing an aglycone intermediate of the formula II

II

wherein R1 represents hydrogen, hydroxyl, or (lower)-alkanoyloxy, in the form of the racemic mixture (7S, 9S; 7R, 9R), which comprises the steps of subjecting a compound of the formula wherein Y represents (lower)alkanoyloxy or in which Z is a ketal residue and W represents a halogen atom to hydrolysis and then separating out the racemic mixture (7S, 9S; 7R, 9R) by conventional procedures.

15. The process according to Claim 14 wherein R1 is hydrogen.

16. The process according to Claim 14 wherein R1 is hydroxyl.

17. The process according to Claim 14 wherein R1 is (lower)-alkanoyloxy.

18. The aglycone intermediate of the formula II, as shown in Claim 14, II

in the form of the racemic mixture (7S, 9S; 7R, 3R) when-ever prepared by the process of Claim 14 or by an obvious chemical equivalent thereof.

19. The aglycone intermediate of the formula II as shown in Claim 18, wherein R1 represents hydrogen, in the form of the racemic mixture (7S, 9S; 7R, 9R) when-ever prepared by the process of Claim 15 or by an obvious chemical equivalent thereof.

20. The aglycone intermediate of the formula II as shown in Claim 18, wherein R1 represents hydroxyl, in the form of the racemic mixture (7S, 9S; 7R, 9R) when-ever prepared by the process of Claim 16 or by an obvious chemical equivalent thereof.

21. The aglycone intermediate of the formula II as shown in Claim 18, wherein R1 represents (lower)alkanoyloxy, in the form of the racemic mixture (7S, 9S; 7R, 9R) when-ever prepared by the process of Claim 17 or by an obvious chemical equivalent thereof.