AP902A

AP902A - Furan and thiophenenecarbothioamide derivatives, their preparation and their use as inhibitors of the replication of HIV-1 and HIV-1 mutants

Info

Publication number: AP902A
Application number: APAP/P/1998/001245A
Authority: AP
Inventors: Walter Gerhard Brouwer; Ewa Maria Osika; Benjamin James Pierce
Original assignee: Uniroyal Chem Co Inc; Uniroyal Chemical Co /Uniroyal Chemical Cie
Priority date: 1995-11-30
Filing date: 1996-11-15
Publication date: 2000-11-23
Also published as: JP3027771B2; PT874839E; EP0874839B1; WO1997019940A1; TW448169B; EP0874839A1; DE69623819D1; AU704086B2; KR19990071682A; CA2237194A1; ATE224382T1; HK1016601A1; ZA969490B; JPH11504657A; CN1203596A; KR100505841B1; DE69623819T2; AU1119997A; CA2237194C; DK0874839T3

Abstract

Compounds of formula (I), wherein X is O or S. The compounds of this invention are useful for the inhibition of the replication of Human Immunod-eficiency Virus-1 (HIV-1) and reverse transcriptase (RT) mutants thereof, in vitro and in vivo. The compounds are useful in the therapeutic or prophylactic treatment of diseases caused by HIV-1 and RT mutants thereof, such as acquired immune deficiency syndrome (AIDS).

Description

FURAN- AND THIOPHENECARBOTHIOAMIDE DERIVATIVES, THEIR PREPARATION AND THEIR USE AS INHIBITORS OF THE REPLICATION OF HIV-1 AND HIV-1 MUTANTS.

Field of the Invention

This invention relates to a compound useful for the inhibition of the replication of HIV-1 and HIV-1 mutant strains. More particularly, this invention relates to methylfuranyl- and methylthienyl-pentenylether derivatives which are inhibitory of the replication of wild-type HIV-1 and HIV-1 reverse transcriptase mutant strains. This invention also relates to a method for the prevention or treatment of HIV-1 infection in a patient which comprises administering to the patient an effective amount of the methylfuranyl- or methylthienylpentenylether derivatives.

Background of the Invention

Various compounds have been described as inhibitors of human immunodeficiency virus type 1 (HIV-1) in vitro and are targeted at the virus-encoded reverse transcriptase (RT), e.g., nevirapine, pyridinone, TIBO,

BHAP, TSAO, and quinoxaline. U.S. Patent No. 5,268,389 describes certain thiocarboxylate ester compounds useful for inhibiting the replication of HIV. The selectivity ' of these compounds for HIV-1 is due to a highly specific interaction with HIV-1 RT.

The rapid emergence of HIV-1 strains resistant to several HIV-l-specific RT inhibitors in cell culture and in AIDS patients has caused concern for further development of these inhibitors in the clinic. For example, HIV-1 strains containing the 100 Leu -> lie mutation in their RT are resistant to TIBO R82913 and

R82150. HIV-1 strains containing the- 138 Glu -> Lys mutation in their RT are resistant to TSAO derivatives.

The 181 Tyr -> Cys mutation in the RT of HIV-1 strains renders the mutant viruses resistant to nevirapine and pyridinone. See, e.g., Balzarini et al, J. Virology

6.7(9): 5353-5359 (1993) (Balzarini I) and Balzarini et

APO 0 0 9 0 2

- 2 al, Virology 192: 246-253 (1993) (Balzarini II¹').

Attempts have been made to combine various HIV-1 RT inhibitors to eliminate virus resistance. See, e.g.,

Balzarini I.

It is the purpose of this invention to provide compounds which, by themselves, can inhibit or suppress the emergence of wild-type HIV-1 and HIV-1 RT mutant strains. It is also the purpose of this invention to provide a method of preventing or treating HIV-1 infections by administration of such compounds.

Summary of the Invention

This invention relates to a compound of the formula

ch₃ ( ι ) wherein X is 0 or S.

The compounds of this invention are useful for the inhibition of the replication of Human Immunodeficiency Virus-1 (HIV-1) and reverse transcriptase (RT) mutants thereof, in vitro and in vivo. The compounds are useful in the therapeutic or prophylactic treatment of diseases caused by HIV-1 and RT mutants thereof, such as acquired immune deficiency syndrome (AIDS).

This invention additionally relates to a pharmaceutical composition comprising a therapeutically effective amount of the compound of formula I and a pharmaceutically acceptable carrier.

AP/P/ 98/01245

This invention also relates to a method of treating HIV-1 infection in an afflicted host which comprises administering to the host a therapeutically effective amount of the compound of formula I.

Description of the Invention

Preferably, this invention relates to a compound of the formula

( IB)

AP/I7 98/01245

Method of Synthesis

The compounds of this invention can be prepared by reacting an acid of the formula A-COOH wherein A is

X with aniline derivative of the formula

ch₃

The acid, A-COOH, is first converted to its acid chloride and then treated with the aniline derivative and an acid scavenger, in a suitable solvent, to form an amide. The acid scavenger can be an organic base, such as pyridine, or an alkali metal hydroxide, carbonate or bicarbonate, such as sodium bicarbonate. Suitable solvents for this step include methylene chloride, diethyl ether, ethyl acetate, or the like. The resultant amide is then reacted with a thionylating agent such as, e.g.,

Lawesson's reagent or phosphorus pentasulfide, in the presence of an acid scavenger, e.g., pyridine or sodium bicarbonate, in an appropriate solvent. Heat is usually applied to complete the thionylation reaction. Suitable solvents for the thionylation reaction include toluene, xylene, DME, and the like.

AP/P/ 9 8/01245

[S] = Thionylating Agent

A second method for making the compounds of this 35 invention is the metallation of a bromo compound of the formula A-Br with n-butyl lithium, at a temperature of -75°C to -80°C. The resultant lithium complex is then

APO π ο 9 Ο 2

- 5 reacted with an isothiocyanate of the formula

CH₃

The compounds of this invention (carbothioamides) are then produced directly after acidification.

Comparative Compounds 1-117 can be prepared in a similar manner using the two methods described above. Typically, the preparation of the comparative compounds involves the formation of the amide and subsequent thionylation with Lawesson's reagent or phosphorus pentasulfide .

AP/P/ 9 8/01245 (i) Base (ii) Lawesson's reagent or P₂S₅ with pyridine or sodium bicarbonate

A, Z and B are as defined in Table 1.

The compounds of the present invention can be administered orally, parenterally, sublingually, by inhalation spray, rectally, or topically in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants, and

AP Ο Ο Ο 9 Ο 2

vehicles. Pharmaceutically acceptable carriers, adjuvants and vehicles useful in the composition of this invention can be found in standard pharmaceutical texts such as, e.g., Remington's Pharmaceutical Sciences, 16th Edition, Mack Publishing Company, Easton, PA (1980).

The therapeutically effective amount of the compounds of this invention that can be combined with the pharmaceutically acceptable carrier to produce a single dosage form will vary depending upon the age and condition of the host treated and the particular mode of administration. In general, the compounds of this invention are most desirably administered at a concentration level that will generally afford antivirally effective results without causing any harmful or deleterious side effects.

While the compounds of this invention can be administered as the sole active pharmaceutical agents, the compounds can also be used in combination with one or more other pharmaceutical agents which are not deleterious to the activity of the compounds of this invention or whose combination with the compounds will not have a deleterious effect on the host treated.

AP/P/ 98/01245

The following examples are provided to illustrate the present invention.

EXAMPLES

Materials and Methods

Test compounds (A) Compound IA was prepared as follows ·.

APO Ο Ο 9 Ο 2 (METHOD I)

Step 1

Preparation of 2-chloro-5-nitrophenol

2-Amino-5-nitrophenol (65.6g) was added to 36% hydrochloric acid (200 mL) at 0 to 5°C with stirring. A solution of sodium nitrite (33.25g) in water (75 mL) was added dropwise over 1.5 hours, after which this first reaction mixture was held at this temperature for a further 1 hour. Excess nitrous acid was decomposed with sulfamic acid (1.5g), added in portions. The first reaction mixture was then added in portions to a stirred suspension of copper (I) chloride (8.5g) in 20% hydrochloric acid (50 mL). Considerable foaming occurred. After the addition, the resultant second reaction mixture was stirred for 1 hour. A precipitate from the second reaction mixture was then collected on a filter, washed with water and dried to give 63.2g of brown solid. This solid was then refluxed with 1.5g of activated carbon in methanol (500 mL) for 15 minutes; filtered through celite; and evaporated, to give 60.8g of brown 2-chloro-5-nitrophenol. '

AP/P/ 9 8 / 0 1 2 4 5

Step 2

Preparation of l-chloro-2-(3-methyl-2-butenyloxy)-4nitrobenzene

A reaction mixture of 2-chloro-5-nitrophenol (50.6g), anhydrous potassium carbonate (44.5g), tetrabutylammonium bromide (4.7g) and 4-bromo-2-methyl-2butene (53.3g, 90%) in methyl ethyl ketone (263 mL) was stirred at ambient temperature overnight. TLC of the reaction mixture showed traces of phenol remaining. Additional prenyl bromide (1 mL) was then added to the reaction mixture and stirred for 2 hours. The solvent was then removed from the reaction mixture. The residue of the reaction mixture was then treated with water and

AP000902

- 8 extracted into diethyl ether. The extract was washed with 2N sodium hydroxide and water, dried (MgSOj , filtered, and evaporated, to leave a brown solid which was recrystallised from ethyl acetate/isopropyl alcohol to give l-chloro-2-(3-methyl-2-butenyloxy)-4nitrobenzene, 49.5g, a beige solid, a single spot on TLC (ethyl acetate : hexane, 20:80) .

Step 3

Preparation of 4-chloro-3 - (3-methyl-2butenyloxy)benzenamine

To a refluxing well-stirred suspension of iron powder (19.6g, 100 mesh) in ethanol (60 mL), water (13.4 mL) and 36% hydrochloric acid (1.4 mL) was added 1chloro-2-(3-methyl-2-butenyloxy)-4-nitrobenzene (24g) in portions over 15-30 minutes. After 3 hours thin-layer chromotography (TLC) (ethyl acetate:hexane, 40:60) showed no substrate. The reaction mixture was then filtered hot and the iron oxide filter cake was washed with hot ethanol. The combined ethanol washes were evaporated to produce a residue. The residue was taken up in diethyl ether, washe*d with aqueous bicarbonate and water, dried (MgSO₄) , filtered, and evaporated, to give 4-chlopo-3-(325 methyl-2-butenyloxy)benzenamine, 19.8g of light brown oil.

AP/P/ 98/01245

Step 4

0 Preparation of 2-methyl-3 - furoyl chloride

A first reaction mixture of chloroacetaldehyde dimethylacetal (300g), water (400 mL) and 36% hydrochloric acid (40 mL) was stirred and brought to reflux. When the first reaction mixture became homogenous, it was cooled and added to a stirred solution of ethyl acetoacetate (260g) and pyridine (500 mL) and left stirring at ambient temperature for 72 hours, to

APO 0 0 9 ο 2 produce a second reaction mixture. The organic layer was then separated from the second reaction mixture and the aqueous layer was diluted with water and then extracted with methylene chloride. The combined organics were washed with 2N hydrochloric acid, followed by removal of the solvent. The residue was treated with a solution of sodium hydroxide (80g) in water (700 mL·) and ethanol (100 mL), to produce a third reaction mixture. After refluxing for 1 hour the third reaction mixture was poured into ice/water and acidified with hydrochloric acid. A cream colored precipitate formed. This precipitate was collected on a filter, washed with water and dried to give 2-methyl-3-furancarboxylic acid, 180g. lOOg of the 2-methyl-3-furancarboxylic acid was added in portions to thionyl chloride (500 mL) and refluxed for 3 hours. Excess thionyl chloride was then distilled off to produce a residue. The residue was distilled using a water pump, to give 2-methyl-3-furanylcarboxylic chloride, bp. 62°C, lOOg.

Step 5 '

Preparation of N-[4-chloro-3-(3-methyl-2- :_t butenyloxy)phenyl]-2-methyl-3 -furancarboxamide

A first solution of 2-methyl-3-furancarboxylic chloride (I3.57g) in methylene chloride (94 mL) was stirred in an ice/salt bath. A second solution of 4chloro-3-(3-methyl-2-butenyloxy)benzenamine (19.8g), triethylamine (14 mL) in methylene chloride (94 mL) was added to the first solution at such a rate that the temperature was maintained at -5°C to 0°C. When the addition was complete, the resultant reaction mixture was stirred to ambient temperature and left stirring for 15 hours. Water was added to the reaction mixture and then the organic layer separated and washed successively with water, dilute hydrochloric acid, water, aqueous sodium bicarbonate, and water. After drying (MgSO₄) , the solvent

AP/P/ 9 8/01245

ΑΡΟ Ο Ο 9 Ο 2

- 10 was removed from the washed organic layer and the residue was recrystallised from isopropyl alcohol to give N-[4chloro-3 - (3-methyl- 2-butenyloxy) phenyl-2-methyl- 3 furancarboxamide, a white solid, 17g. A second crop of beige crystals, 7.6g, was also obtained.

Step 6

Preparation of N- [4-chloro-3-(3-methy-2-butenyloxy)phenyl] -2 -methyl-3 - furancarbothioamide (Compound IA)

A reaction mixture of N-[4-chloro-3-(3-methyl-2butenyloxy)phenyl]-2-methyl-3-furancarboxamide (4g), sodium bicarbonate (7.4g) and Lawesson's reagent (3.6g) in toluene (168 mL) was gradually heated to 85°C over 1 1/2 hours and then held at this temperature for a further 2 1/2 hours. The reaction mixture was then cooled and filtered through a plug of neutral aluminum oxide and eluted with ether: petroleum ether (1:1). Evaporation of the solvent gave the product, N-[4-chloro-3-(3-methyl-2butenyloxy)phenyl]-2-methyl-3-furancarbothioamide, 2.6g.

(B) Compound IB was prepared as follows:

o (METHOD II)

Step 1

Preparation of 3,5-dibromo-2-methylthiophene

2-Methylthiophene (98g) in dioxane (500 mL) was stirred while bromine (320g) in dioxane (2L) was added dropwise over 7.5 hours and then left to stand at ambient temperature overnight. The reaction mixture was then heated to reflux for 3 hours, poured into water (4L), and extracted with ether. The ether extract was washed with aqueous bicarbonate and then water, and dried (MgSO₄) . Evaporation left an oil which was distilled to give 3,5dibromo-3-methylthiophene, bp. 98°C, 234.6g.

AP/F/ S 8 / 0 1 2 4 5

ΑΡΠ009θ2

- 11 10

Step 2

Preparation of 3-bromo-2-methvlthiophene

Magnesium turnings (5g) were covered with tetrahydrofuran (THF) (25 mL), and with stirring, treated with 3,5-dibromo-2-methylthiophene (5g). An exotherm developed. The temperature of the reaction mixture was maintained at 35°C + 5°C with cooling. Additional 3,5dibromo-2-methylthiophene (47g) was added dropwise to the reaction mixture while the temperature of the reaction mixture was maintained in range stated above. Towards the end of the addition, the temperature was allowed to rise to 40°C. Most of the magnesium had reacted. When the reaction exotherm ceased, the temperature of the reaction mixture was raised from 40°C to 50°C by external heating and maintained at 50°C for 1 hour. The reaction mixture was then poured slowly onto vigorously-stirred ice/water/dilute hydrochloric acid, to produce a second mixture. The second mixture was extracted, into ether, washed with water, dried (MgSOj , and evaporated, to leave a liquid which was distilled at the water pump to give 3bromo-2-methylthiophene, a clear liquid, bp. 62-65°C,

2.6g

AP/P/ 9 8/01245

Step 3

Preparation of l-chloro-2-(3-metbvl-2-butenyloxy)-4isothiocyanatobenzene

1-Chloro-2-(3-methyl-2-butenyloxy)benzenamine (19.9g) (c.f., Method I, Step 3) was dissolved in methylene chloride (75mL), to produce a first solution. With vigorous stirring, the first solution was then covered with ice/water (250 mL). A second solution of thiophosgene (7.3 mL) in methylene chloride (25 mL) was then added dropwise to the first solution. The resultant reaction mixture was then kept cool with an ice bath. A solid precipitated from the reaction mixture, but with time, the solid reacted further and dissolved in the

AP Ο Ο ο 9 Ο 2

methylene chloride. The reaction mixture was then allowed to come to ambient temperature by which time the reaction was complete. The organics were then separated from the reaction mixture, washed with water, dried and evaporated under reduced pressure (a caustic scrubber was used to absorb any excess thiophosgene), to produce a dark grey solid. The solid was purified by dissolving in commercial hexanes, passing the dissolved solid through a column of silica gel (minimum 3.5 cm diam. x 6 cm length), and then eluting the column with hexanes.

Evaporation of the hexanes solution gave a white solid, l-chloro-2-(3-methyl-2-butenyloxy)-4-isothiocyanatobenzene, 22g, a single spot on TLC (hexane).

m

CM

Step 4 v—

Preparation of N-[4-chloro-3-¢3-methvl-2-butenyloxy)- O phenyl]-2-methyl-3thiophenecarbothioamide (Compound #IB)

Under an atmosphere of nitrogen, a solution of 3- CO bromo-2-methylthiophene (5.5g) in dry ether (25 mL) was cooled in an acetone/dry ice bath. n-Butyl lithium (10 CX mL, 2M in hexanes) was added dropwise to the solution (X such that the temperature of the resultant first reaction mixture did not rise by more than 2°C. After this addition, the first reaction mixture was stirred at -75°C for 1.5 hours. l-Chloro-2-(3-methyl-2-butenyloxy)-4isothiocyanatobenzene (6.2g) in dry ether (25 mL) was added dropwise to the first reaction mixture at such a rate that the temperature of the resultant second reaction mixture did not rise by more than 2°C (1.25 hrs). After this addition, the second reaction mixture was allowed to slowly come to ambient temperature over 2 hours. The second reaction mixture was then treated with ice/water/dilute hydrochloric acid/ether. The organics were then separated from the second, reaction mixture.

The organics were then washed with water, dried (MgSO„) , and the solvents removed. A viscous yellow oil remained.

APO 0 0 9 0 2

- 13 A yellow solid was obtained when this oil was stirred with commercial hexanes. The yellow solid was collected on a filter, 6.2g, and recrystallised from cyclohexane/ether to give N-[4-chloro-3-(3-methyl-25 butenyloxy)phenyl-2-methyl-3-thiophenecarbothioamide, a yellow solid, mp 118-120°C (uncorrected), 4.7g.

Compounds IA and IB gave satisfactory I.R. and NMR 10 spectra.

AP/P/ 9 8/01245

V

APO ο ο 9 Ο 2

Table 1

ζ

Β

C3

Cmpd.	Y	z	A
1	S	Cl	2-methyl-3thienyl
2	tl	II	2-methyl-3furanyl
3	II	tt	II
4	II	-och₃	II
5	II	Cl	6-methyl-4,5dihydro-2Hpyran-5-yl
6	II	It	2-methyl-3furany1
7	It	II	2-methyl-4,5-dihydro-3-furanyl
8	II	II	2-methyl-3thienyl
9	II	It	II
10	II	It	1,2-dimethyl-3 pyrrolyl
11	II	tl	1-methyl-3pyrrolyl
12	II	II	2-methyl-3thienyl
13	o	II	5,6-dihydro-2ethyl-1,4oxathiin-3-yl
14	s	II	2-methyl-3thienyl
15	0	II	5,6-dihydro-2ethyl-1,4oxathiin-3-yl
16	s	11	2-methyl-3thienyl
17	II	II	2-methyl-3furanyl

-OCH₂CH=CHCH₃

II

-CH=N-O-C(CH₃)₃

-ch=n-o-ch₃

-CH=N-O-C(CH₃) ₃

-s-ch₂ch=chch₃

-ch=n-o-c(ch₃)₃

-s-ch₂ch=chch₃

I I

-c

-Ο-ΛΛ

-CH=N-O-C (CH₃) j

-O-CH₂CH₂CH(CH₃) j .ο-Λύ

-O-CH₂-C (O) -o-c (ch₃) ₃

-CH=N-O-C (CH₃) ₃

-C(0)-O-CH(CH₃)j

-S-CH₂-C (0) -O-C (CHj) ₃

AP/P/ 9 8/01245

APO 0 0 9 0 2

- 15 Table 1 (continued)

B

Ctnpd.Y_Z

O Cl

S

O

Cl

S CN

Cl

O

S

II II

5,6-dihydro-2methyl-1,4oxathiin-3-yl

1,2 -dimethyl-3pyrrolyl

2-methyl-3thienyl

5,6-dihydro-2methyl-1,4oxathiin-3-yl

2-methyl-3thienyl

6-methyl-4,5dihydro-2Hpyran-5-yl

2-methyl-3thienyl

2-methyl-3furanyl

2-methyl-3thienyl

-CH=N-O-C (CH₃) ₃

-O-CH₂CH=CHCH₃

-S-CHj-C(O) -O-C(CH₃) ₃

-C-O-CH

CH(CH₃)₂

II CH(CH₃)₂

C-O-CH

CH(CH₃)₂ .0-0-/3

-O-CH₂-C{C1)=CH,

-O-CH₂CH=CH(CH₃) ·

O-CH₂-^2^

-0CH₂C(CHj)₃

AP/P/ 9 8 / 0 1 2 A 5

AP Ο Ο Ο 9 Ο 2

Table 1 {continued)

Cmpd. Υ_Ζ

Ο C1

2-methyl-3furanyl

2-methyl-3 thienyl

-OCH₂CH=C{CH₃)₂

-C(0)-O-CH{CH₃)₂

II CH(CH₃)₂

-C-O-CH

CH(CH₃)₂

O

2-methyl-3 furanyl

2-(methylthio)3-thienyl

2-methyl-3furanyl

1,2-dimethyl-3pyrrolyl

2-methyl-3furanyl

II

5,6-dihydro-2 methyl-1,4oxathiin-3-yl

-O-CH₂CH₂CH(CH₃) ₂-S-CH₂CH=CHCH₃

-s-ch₂ch=chch₃

-CH=N-O-C(CH₃)₃

-O-CH₂CH₂CH(CH₃)₂

-CH=N-O-C (CHj) ₃

-O-CH₂C(CH₃)₃

-o-ch₂ch=chch₃

-O-CH₂CH₂CH(CH₃) 2

AP/P/ 9 8/01245

2-methyl-3thienyl

-O-CH

S

O

5,6-dihydro-2ethyl-1,4 oxathiin-3-yl

2-methyl-3furanyl

2-methyl-3thienyl

-O-CH₂CH=CHCH₃

-o-ch₂ch₂ch{ch₃) ₂

-O-CH₂CH=CHCH₃

AP Ο Ο Ο 9 Ο 2

- 17 Table 1 (continued)

Ν'

	Cmpd.	Υ	ζ	A	B
	46	S	Cl	2-methyl-3 furanyl	-O-C(O) -O-CH(CH₃)₂
	47	0	-och₃	11	-ch=n-o-ch₃
	48	0	Cl	II	-C(O) -O-CH(CH₃)₂
15	49	S	II	II	II
	50	Ο	π	II	-C(O) -O-CH₂C(CH₃) =ch₂
	51	S	II	II	II
	52	0	ft	II	-CH₂-O-C(CH₃) J
	53	S	II	II	It
20	54	0	11	II	-C(O) -O-CH(CH₃) cyclo-C₃H₅
	55	S	II	It	II
	56	0	-OCHj	II	-CHj-C(O) -O-CH(CH₃)₂
	57	S	II	tr	II
	58	0	Cl	2-methyl-3- thienyl
25	59	S	II	II	II *
	60	0	' II	II	-CH=N-O-C(CH₃) ₃
	61	S	II	II	II
	62	0	It	Phenyl	-C(O) -O-CH{CH₃)₂
	63	S	II	II	II
30	64	0	II	2 -fluorophenyl
	65	S	11	II	II
	66	0	11	2 -aminophenyl	' -C(O) -O-CH(C₂H₅)₂
	67	S	II	11	II
	- 68	0	It	2-methoxyphenyl	-C-O^
35	69	S	II	(1	II

Cmpd. Υ

AP ο π Ο 9 Ο 2

- 18 Table 1 (continued)

Ο I I

	70	Ο	C1	2-methylphenyl
	71	S	II	II	11
	72	S	C1	2-methyl-3furanyl	-ch=n-o-ch₃
15	73	II	-OCHj	It	-o-ch₂-ch=ch₂
	74	II	-CN	II	II
	75	Η	-ch₃	It	-C(O) -O-CHjCH₃
	76	II	Cl	3-methyl-2thienyl	-o-ch₂-ch=ch₃
	77	II	-CN	11	II
20	78	II	-och₃	It	II
	79	II	ci	3-thienyl	.L-o
	80	II	II	2 -chlorophenyl	II
	81	It	II	2-methoxyphenyl	II
	82	It	II	phenyl	II
25	83	η	II	2-methyl-3furanyl	It
	84	II	II	2-methylphenyl	II
	85	II	II	2 -aminophenyl	-C(O)-O-CH(CH₃) ₃
	86	IJ	tl	2 -furanyl	It
	87	η	tl	3-methyl-2- furanyl	II
30	88	II	It	3-methyl-2thienyl	II
	89	II	II	2-thienyl	II
	90	η	It	2-methoxyphenyl	II
	91	II	ft	2 -methylphenyl	II
	92	II	II	3-thienyl	II
35	93	II	II	phenyl	-CH=N-O-C(CH₃) ₃

ΑΡ/Ρ/ 9 8 / 0 1 2 4 5

Cmpd.

Cl

100

101

102

103

104

105

106

107

108

109

110

ΑΡΟ 0 0 90 2

- 19 Table 1 (continued)

3-methyl-2furanyl

3-methyl-2 thienyl phenyl

2-methyl-3furanyl •CH=NOCH(CH₃).

2-fluorophenyl

2- pyrrolyl

1- methyl-2pyrrolyl

3- methyl-2thienyl

2- metlprl-3furanyl

II PH(CH₃)₂

-C-O-CH

CH(CH₃)₂

-C (O) -O-CH₂CH(CH₃) ₃-C (Ο) -O-CH₂CH=CH₂-C(O) -0-CH₂-cyclo-C₃H_s-C(O)-O-CH₂-CF₃-C (Ο) -O-CH (CH₂CH₃) ₂-C (Ο) -O-CH₂CH(CH₂CH₃) J •oO o CH,

II I ³- C-0-C-CH=CH,

CH,

AP/P/ 9 8/01245

APO 0 0 9 0 2 }

- 20 10

Cmpd. Y_Z

111 S Cl

112

113

114

115

116

117

Table 1 (continued)

B

2-methyl-3furanyl

It

II

- CH=N - O - CH₂ CH=CH₂

-CH=N-O-CH₂CsCH

-CH--N-0-O

-CH=N-O-C(CII₃) 3 -O-CHj-C(O) -O-C(CHj)₃-S-CHj-C(O) -O-CH₂CH₃

-o-ch₂-c=ch

AP/P/ 9 8 / 0 1 2 4 5

AP 0 0 0 9 0

- 21 Cells and Viruses

CEM cells were obtained from the American Tissue Cell Culture Collection (Rockville, Md.). HIV-1(III_B) was originally obtained from the culture supernatant of persistently HIV-l-infected H9 cells and was provided by R.C. Gallo and M. Popovic (National Cancer Institute, National Institutes of Health, Bethesda, MD). The selection and characterization of the HIV-1 RT mutant strains were done as follows: HIV-l/lOO-Ile (100-Ile) was selected for resistance against TIBO R82150 as described in Balzarini et al, Virology 192: 246-253 (1993); HIV-l/103-Asn (103-Asn) was selected for resistance against TIBO R82913 as described in Balzarini et al, Virology 192: 246-253 (1993); HIV-l/lO6-Ala (106-Ala) was selected for resistance against nevirapine as described in Balzarini et al, J. Virol. 67: 5353-5359 (1993); HIV-l/Lys-138 (Lys-138) was selected for resistance against TSAO-m³T as described in Balzarini et al, Virology 192: 246-253 (1993) and Balzarini et al,

Proc. Nat. Acad. Sci. USA 90: 6952-6956 (1993); HIV^Al/l81-Cys (181-Cys) was selected for resistance against pyridinone L-697,661 as described in Balzarini et al·, Virology 192: 246-253 (1993); and HIV-1/188-His (188- 25 His) was selected for resistance against HEPT as described in Balzarini et al, Mol. Pharmocol. 44: 694-701 (1993). 188-His was then further converted to HIV-1/188Leu(188-Leu) upon further passage in cell culture in the absence of the HEPT.

Antiviral activity of the test compounds in cell cultures

CEM cells were suspended at =300,000 cells per ml of culture medium and infected with approximately 100 CCID₅₀ (CCID_5c being the 50% cell culture infective dose) of HIV-1 (III_B) or one of the HIV-1 RT mutant strains described above. Then 100 μΐ of the infected cell

P/P/ 9 8/01245

APO 0 Ο 9 Ο 2 ί ^; )

- 22 suspensions was added to 200 μΐ microtiter plate wells containing 100 μΐ of appropriate serial (5-fold) dilutions of the test compounds. The inhibitiory effect of the test compounds on HIV-1 induced syncytium formation in CEM cells was examined microscopically on day 4 post infection. The 50% effective concentration (EC₅₀) was defined as the test compound concentration that inhibits syncytium formation in the HIV-1-infected cell cultures by 50%.

Results

As seen below in Tables 2A and 2B, compounds IA and IB had extremely low, and virtually similar EC₅₀ values against HIV-1(III_B) (designated as ir>

C\J

O

OO o>

o.

ΑΡ ο (I Ο 9 Ο 2

- 23 Table 2Α

Antiviral Activity⁰

Cmpd.	EC_5o (/7g/ml)·	cc₆₀ Ct/g/ml)^b
WT	100-lle	103-Asn	106-Ala	138-Lys	181-Cys	188-Leu
IA	0.003	0.003	0.006	0.005	0.005	0.011	0.50	>100
IB	0.003	0.004	0.004	0.005	0.005	0.005	0.60	5.8

* 50% effective concentration (i.e., compound concentration required to inhibit virus-induced cytopathicity by 50%) ^b 50% cytostatic concentration (i.e., compound concentration required to inhibit OEM cell proliferation by 50%) ‘ Data are the averages of 2 to 3 independent experiments

AP/P/ 9 8/01245

AP Ο Ο Ο 9 Ο 2

- 24 Table 2Β

Antiviral Activity of Comparative Compounds⁰

Cmpd.	EC₅₀ {/vg/m!)·	CC₅₀ (t/g/ml)^b
WT	100-Ile	103-Asn	106-Ala	138-Lys	181-Cys	188-Leu
1	0.0055	0.048	0.048	0.023	0.018	0.03	0.65	5.9
2	0.006	0.048	0.023	0.04	0.018	0.075	0.65	9.0
3	0.0065	0.057	0.23	0.03	0.045	0.08	0.91	5.8
4	0.007	0.22	0.4	0.22	0.039	0.59	0.93	>100
5	0.013	0.055	0.08	0.025	0.04	0.025	>2	10
6	0.0041	0.037	0.03	0.018	0.03	0.045	1.0	4.7
7	0.0065	0.065	0.058	0.03	0.04	0.045	1.2	11
8	0.0055	0.032	0.023	0.023	0.025	0.025	ϊ>2.0	5.1
9	0.0032	0.057	0.13	0.03	0.02	0.032	>2	6.0
10	0.02	0.65	1.27	0.23	0.09	0.033	>2	6.5
11	0.029	0.13	0.17	0.1	0.13	0.045	>2	13
12	0.007	0.035	0.035	0.08	0.035	0.07	>2.0	>100
13	0.08	0.26	0.38	0.17	0.25	0.07	>2	11.4
14	0.067	0.16	0.12	0.1	0.24	0.087	>2	2.1
> 15	0.03	0.06	0.065	0.06	0.07	0.09	>2	c 11.4
16	0.0041	0.15	0.33	0.077	0.03	0.1	Ξ>2.0 i	5.9
17	0.22	' 0.49	0.73	0.3	0.18	0.45	>2	7.0
18	0.014	0.33	0.33	0.15	0.22	0.13	>2	8.5
19	0.17	0.85	>2.0	0.4	0.6	0.13	>2	5.2
20	0.13	0.34	0.44	0.15	0.43	0.14	>2	5.4
21	0.008	0.3	0.61	0.23	0.11	0.17	>2	6.0
22	0.08	0.48	0.65	0.26	0.55	0.18	>2	5.8
23	0.03	0.7	>2.0	0.22	0.73	0.27	>2	5.8

APO Ο Ο 9 ο 2

- 25 Table 2Β

Antiviral Activity of Comparative Compounds^c

Cmpd.	EC_sotpg/m1)-	cc₅₀
WT	100-lle	103-Asn	106-Ala	138-Lys	181-Cys	188-Leu	(yg/ml)^b
24	0.13	0.48	0.6	0.5	0.5	0.33	>2	9.7
25	0.03	0.2	0.4	0.2	0.13	0.44	>2	3.3
26	0.033	0.33	0.33	0.09	0.50	>2.0	>2	5.3
27	0.01	0.36	0.5	0.11	0.15	0.46	>2	6.0
28	0.029	0.22	0.33	0.23	0.11	0.5	>2	4.3
29	0.14	0.5	0.43	0.5	0.05	0.55	>2	6.2
30	0.015	1.27	1.67	0.34	0.23	0.65	>2	>100
31	0.09	>2.0	>2.0	0.6	0.93	0.73	>2	5.5
32	0.16	>2.0	>2.0	>2.0	>2.0	0.95	>2	4.1
33	0.063	0.95	1.0	0.65	0.62	1.07	>2	7.0
34	0.08	0.65	0.8	0.28	0.5	1.2	>2	4.8
35	0.16	1.5	0.55	0.8	0.7	1.6	>2	5.9
36	0.18	1.23	1.2	0.85	0.85	1.6	>2	7.8
37	0.16	>2.0	>2.0	1.3	1.25	1.73	>2	5.0
38	0.08	>2.0	>2.0	0.7	>2.0	>2	>2	5.5
39	0.15	.1-4	0.7	>2	0.5	>2.0	>2	4.1.
40	0.24	1.4	>2.0	>2	1.2	>2	>2	5.8
41	0.28	>2.0	>2.0	>2.0	>2.0	>2.0	>2	>100
42	0.36	>2.0	>2.0	>2.0	>2.0	>2.0	>2	5.0
43	0.65	1.2	1.3	4.5	1.3	>2	>2	11
44	0.0073	0.035	0.08	0.06	0.03	0.08	>10	>100

AP/P/ 98/01245

APO 0 0 9 0 2

- 26 Table 2B (continued)

Antiviral Activity of Comparative Compounds^c

Cmpd.	EC_5o (//g/ml)*	cc_so (/yg/ml)^b
WT	100-lle	103-Asn	106-Ala	138-Lys	181-Cys	188-Lbu
45	0.15	0.6	0.95	0.85	0.65	0.5	>10.0	87
46	0.37	4.5	4.0	3.0	0.6	5.5	>2	25
47	0.23	>10	>10	>10	>10	>10	>10	>100
48	0.45	6.5	-	6.5	3.5	6.0	—	3.8
49	0.007	0.6	-	0.6	0.6	0.2	—	5.2
50	0.15	5.0	-	1.0	1.0	1.9	-	3.7
51	0.006	0.8	-	0.03	0.03	0.10	—	6.7
52	0.70	8.5	-	5.5	4.0	>10	-	6.2
53	0.03	0.85	-	0.29	0.08	0.55	-	'18
54	0.6	>10	-	5.0	>10	1.0	—	44
55	0.009	0.65	-	0.08	0.18	0.11	—	17
56	0.7	>10	-	6.5	>10	>10	-	52
57	0.009	1.3	-	0.2	0.08	0.50	-	>100
58	0.19	5.5	-	7.0	5-0	5.0	-	6.5
59	0.09	0.65	-	0.60	0.40	0.35	-	15
60	0.23	4.5	-	1.6	1.57	3.0		14
61	0.01	0.2	-	0.2	0.05	0.2	-	12
62	0.4	3.5	-	5.0	5.0	5.5	-	6.2
63	0.05	4.5	-	5.0	2.2	5.0	-	4.3
64	0.6	>10	-	2.0	4.0	5.0	-	33
65	0.04	0.3	-	0.26	0.09	0.25	-	19
66	0.6	>10	-	9.0	>10	>10	-	25

AP/P/ 9 8 / 0 1 2 4 5

AP Ο Ο Ο 9 Ο 2

- 27 Table 2Β (continued)

Antiviral Activity of Comparative Compounds'³

Cmpd.	EC_5o Oug/ml)*	cc₅„ (pg/ml)^b
WT	100-lle	103· Asn	105-Ala	138-Lys	181-Cys	188-Leu
67	0.75	>10	-	>10	5.5	>10	-	29
68	0.45	5.0	-	8.5	5.5	8.5	—	>100
69	0.04	0.65	-	0.65	0.55	3.5	—	10
70	0.19	2.5	-	6.5	5.5	2.5	-	5.2
71	0.049	0.6	-	3.0	3.0	1.6	—	4.0
72	0.03	2.3	-	0.5	0.085	1.6	—	24
73	0.017	2.9	-	0.80	0.09	0.85	—	>100
74	0.015	6.0	-	0.45	0.09	3.0	-	>100
75	0.009	0.9	-	0.7	0.35	0.6	-	16
76	0.035	2.7	-	>1	0.73	>1	-	5.3
77	0.045	>1	-	>1	0.55	>1	-	3.6
78	0.045	5.0	-	2.0	1.13	3.3	-	7.1
79	0.03	0.85	-	0.07	0.07	0.08	-	4.1
80	0.50	4.0	-	5.0	4.0	3.0	-	8.3
81	0.04	0.7	-	0.65	0.6	4.5	- „	12
82	0.03	2.0	-	1.0	0.55	0.8	-	5.2 ;
83	0.004	0.17	-	0.057	0.035	0.07	-	9.7
84	0.049	0.6	-	3.0	3.0	1.6	-	4.0
85	0.5	6.0	-	7.5	5.5	5.3	-	6.7
86	0.1	5.0	-	5.0	4.0	5.0	-	7.4
87	0.02	3.0	-	1.0	0.55	0.53	-	15
88	0.01	0.7	-	0.6	0.3	0.27	-	5.2
89	0.09	3.0	-	4.0	1.9	4.0	-	4.9

AP/P/ 9 8/01245

APO 0 0 9 0 2

- 28 10

Table 2B (continued)

Antiviral Activity of Comparative Compounds'^

Cmpd.	EC₅₀(qg/ml)·	cc₅„ Uvg/ml)^b
WT	100-lle	103-Asn	106-Ala	138-Lys	181-Cys	188-Leu
90	0.19	3.5	-	6.0	0.7	5.5	-	27 '
91	0.45	4.5	-	8.5	5.0	7.5	-	5.8
92	0.02	0.4	-	0.75	0.50	0.37	-	4.3
93	0.03	2.0	-	0.10	0.08	0.7	-	9.7
94	0.01	0.75	-	0.50	0.15	0.6	-	3.1
95	0.008	0.65	-	0.08	0.16	0.5	-	13
96	0.05	>1	-	0.75	0.17	> 1	-	9.0
97	0.009	3.5	-	>4	2.0	3.3	-	3.3
98	0.06	>1	-	0.85	0.7	>1	-	7.5
99	0.10	>1	-	>10	>10	>10	-	>100
100	0.03	4.0	-	0.8	0.4	0.65	-	>100
101	0.045	3.0	-	0.2	0.33	0.55	-	£±100
1&2	0.03	0.85	-	0.85	0.08	0.65	-	3.7
103	0.01	0.19	-	0.075	0.06	0.07	-	8.5
104	0.01	0.65	-	0.4	0.2	0.37	--	11
105	0.015	0.4	-	0.2	0.085	0.07		6.7
106	0.007	0.5	-	0.08	0.065	0.085	-	' >100
107	0.04	0.6	-	0.3	0.15	0.3	-	16
108	0.007	0.27	-	0.08	0.07	0.08	-	7.0
109	0.005	0.17	-	0.04	0.035 .	0.045	-	4.3
110	0.04	4.0	-	0.2	0.1	0.5	-	25
111	0.007	0.5	-	0.085	0.077	0.3	-	12
112	0.025	0.75	-	0.35	0.5	0.8	-	10.

in

-<r

CM o

oo o>

ix cC «X

APO Ο Ο 9 Ο 2

- 29 Table 2Β (continued)

Antiviral Activity of Comparative Compounds^c

Cmpd.	EC_5o (χ/g/ml)*	CC₅₀ (/vg/ml)^b
WT	100-lle	103-Asn	106-Ala	138-Lys	181-Cys	188-Leu
113	0.023	0.27	-	0.085	0.15	0.13	-	5.4
114	0.05	0.085	-	0.13	0.075	0.075	-	5.8
115	0.29	0.65	-	0.2	0.77	0.5	—	4.3
116	0.085	1.8	-	5.5	0.6	2.0	—	>100
117	0.06	5.0	-	0.75	0.35	4.0	-

• 50% effective concentration ji.e., compound concentration required to inhibit virus-induced cytopathicity by 50%) ^b 50% cytostatic concentration (i.e., compound concentration required to inhibit CEM cell proliferation by 50%) ' Data are the averages of 2 to 3 independent experiments

Claims

1. A compound of the formula ch₃

10 (') wherein X is O or S.

2. A compound as recited in claim 1 having the 15 formula

AP/P/S 8/0 1 245

3 .

formula

A compound as recited in claim 1 having the

4. A pharmaceutical composition comprising a therapeutically effective amount of the compound as recited in claim 1 and a pharmaceutically acceptable carrier.

ΑΡΟΟ 0 9 0 2

5. A pharmaceutical composition comprising a therapeutically effective amount of the compound as recited in claim 2 and a pharmaceutically acceptable carrier.

6. A pharmaceutical composition comprising a therapeutically effective amount of the compound as recited in claim 3 and a pharmaceutically acceptable carrier.

7. Use of a compound as recited in claim 1 in the manufacture of a medicament to treat HIV-1 infection in an afflicted host.

8. Use of a compound as recited in claim 2 in the manufacture of a medicament to treat HIV-1 infection in an afflicted host.

9. Use of a compound as recited in claim 3 in the manufacture of a medicament to treat HIV-1 infection in an afflicted host.

10. Use of a compound as recited in claim 1 in the manufacture of a medicament to inhibit the replication of HIV-1.

11. Use of a compound as recited in claim 2 in' the

AP/P/ 9 8/ 0 1 2 4 5 manufacture of a medicament to inhibit the replication of HIV-1.

12. Use of a compound as recited in claim 3 in the manufacture of a medicament to inhibit the replication of HIV-1.

13. A substance or composition for use in a method of treating HIV-1 infection in an afflicted host, . said substance or composition comprising the compound as recited in any one of claims 1 to 3, and said method comprising administering to the host a therapeutically effective amount of said substance or composition.

APOΟ ο 902

- 32 14. A substance or composition for use in a method of inhibiting the replication of HIV-1, said substance or composition comprising a compound as recited in any one of claims 1 to 3, and said method comprising contacting the HIV-1 with an effective amount of said substance or composition .