throbber
WO 90/05719
`
`V
`
`.
`
`V
`
`PCT/GB89/01399
`
`50
`
`[4-Hydroxy-2Reisobutyl-BS-(2,4-dimethylphenylthio-
`1
`2 methyl)succinyl]—L+phenylalanine—N-methylamide (1.8g;
`3
`3.7 mmol) and HOBT (0.67g, 12 mmol) were dissolved in
`4
`l:l7DCM/DMF and the mixture cooled to 0°C before adding
`5 WSDCI'(0.86g, 4.5mmol) and NMM (0.45g, 4.5mmol}. The
`6 mixture was stirred at 0°C for 1h to ensure complete
`7
`formation of
`the activated ester. Hydroxylamine
`8
`hydrochloride (0.39g, 5.6mmol) and NMM (0.56g, 5.6mmol)
`9 were dissolvéi
`in DMF
`then this mixture was added
`10
`dropwise to the cooled solution of the activated ester.
`11 After 1h the reaction was poured into ether/water (1:1)
`12 whereupon the desired product precipitated as white
`137'Crystals. These were collected by filtration, further
`14 washed with ether and water,
`then dried under vacuum at
`15
`50°C. This material was repeatedly recrystallised from
`16 methanol/water
`(1:1)
`to remove a trace of the minor
`17‘
`diastereomer (1.08g, 2.2mmol, 58%).
`18
`'
`
`19 m.p. 225°C (dec.)
`20
`
`21 Analysis calculated for CZ7H37N3O4S
`22 Requires: 064.90 H7.46 N8.4l
`23
`Found:
`C65.15 H7.48 N8.40
`
`24
`'a
`25 ' deltaH (250MHz, 06-DMSO) 8.83 (1H, s, NHog), 8.32 (1H,
`26
`d, J = 8H2, CONE), 7.85 (1H, d, J = 6H2, CONflMe), 7.30
`27
`- 6.71 (9H, m, aromatic H), 4.56 (1H, m, CECHZPh), 2.91
`28
`(1H, ,dd,
`J =: 14,4Hz, CHCflzPh),
`2.76 (13,
`dd,
`.J =
`29
`14,10Hz, CHCflzPh), 2.57 (3H, d, J = 4.5Hz, NHCfi3), 2.53
`30
`- 2.38 (2H, m), 2.23 (3H, s, 06H5(cg3)2), 2.13 (3H, s,
`:;
`c6H5(cg3), 1.30 (2H, m), 0.89 (1H, m. chCH(cH3)2),
`
`0.81 (3H, d, J = 6H2, CH(Cfl3)2), and 0.74 (3H, d, J’=
`'
`-
`6Hz,.CH(cg3)2).
`'7
`,
`.
`_
`
`33
`
`.
`
`&
`
`'
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0701
`
`page 0701
`
`

`

`WO 90/05719
`
`, PCT/GBS9/01399
`
`51
`
`Exam le 15
`
`mummsz—I
`k0
`
`10
`11
`12
`
`H
`
`o
`
`NHMe
`
`H
`P“
`
`[4(N-Hydroxyamino-ZR—isobutyl—BS—(acetylthiomethyl)
`13
`succinyl]-L-phenylalanine—N—methylamide (1.0g, 2.4
`14
`15 mmol) was dissolved in 750ml methanol and 350ml pH 7
`16
`buffer added. Left
`to stand overnight and solvent
`
`17
`18
`19
`
`20
`
`removed in vacuo to 2/3 volume,left to crystallise for
`a further two hours. Filtered and dried to give 0.87g
`
`off-white crystals
`
`21 Analysis calculated for C38H56N60882.1.9H20
`22 Requires: C55.34 H6.93 N9.88
`23
`Found:
`€55.44 H7.32 N10.21
`24
`
`25
`26
`
`27
`28
`29
`
`3o
`31
`
`32
`
`33
`
`Example 16
`
`[4—(N—Hydroxyamino)-2R-isobutyl-3s—(3—bromophenyl-
`thiomethyl) succinyl]-L-phenylalanine-N—methylamide
`Ph
`
`0
`
`fiH
`_
`_H *
`N
`
`NHMe
`
`
`
`o
`
`CONHOH
`
`IN“
`
`Br
`
`8
`
`AQUESTIVE EXHIBIT 1004 page 0702
`
`AQUESTIVE EXHIBIT 1004 page 0702
`
`

`

`wo 90/05719
`
`7
`
`'
`
`PCT/GB89/01399
`
`.U‘AL’JMA
`00x10»
`
`11
`
`12
`
`13
`
`,14
`
`15
`
`16
`
`17'
`
`18
`
`19
`
`.20
`
`21
`
`.22
`
`23
`
`24
`
`25
`
`1’26
`
`27
`
`28
`
`29
`
`3o
`
`31
`
`32
`
`33
`
`52
`
`Prepared by the method described in example lg to give
`material with the following characteristics.
`
`m.p. 225 -2299c
`
`[alphajfi = -164.8°
`
`Analy51s calculated for C2 H3ZBrN3O4S
`
`Requires:
`
`€54.40 H5.89 N7.40
`
`Found:
`
`€54.54 H5.86 N7.637
`
`deltaH (250MHz, D6-DMSO) 8.83 (1H, s, NHOfl), 8.35 (1H,
`d, J’: 8H2, CONE), 7.90 (1H, q, J = 6Hz,,CON§Me), 7.35
`
`- 6.87 (9H, m, aromatic H), 4.64 (1H, m, CgCHzPh), 2.94
`
`(1H, dd, J = 14,4Hz, CHCflZPh), 2.76 (1H, t, J = 13Hz,
`'cncgzph) 2.60 (3H, d, J : 5H2, NHcg3), 2.55 - 2.35 (23,'
`m, c325), 2.15 (1H, t, J = lOHz, chO), 2.01 (1H, d, J
`,4 11.5Hz,
`CECO),
`1.37 (2H, m),
`0.88 (1H, m,
`CEZCH(CH3)2), 0.81 (3H, d, JV: 6H2, CH(c33)2), and 0.74
`(3H, d,J = 6HZ,CH(Cg3)2)..
`'
`
`deltac
`(63.9MHz,7DS~DMSO) 173.0, 171.0, 168.8, 139.8,
`l38.0,:130.5, 129.0, 128.5, 127.5, 125.8, 125.5, 54.2,
`46.0, 45.5, 38.0, 31.5, 25.5, 25.2, 24.7, and 21.0.
`
`Example 17
`
`[4-(N-Hydroxyamino)-2Rfiisobutyl—3s-(3—chlorophenylthio-
`
`methyl) succinyl J —L-phenylalanine-N-methylamide
`'
`,
`rPh
`
`O,
`
`u)
`
`\\H
`°
`
`NHMe
`
`-
`
`H,
`N
`
`
`
`
`
`,
`CONHOH
`
`O
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0703
`
`page 0703
`
`

`

`WO 90/05719
`
`.
`
`PCT/GB89/01399
`
`'
`
`53
`
`1
`
`Prepared by the method described in example lg to give
`
`2 material with the following characteristics.
`
`m.p. 231—2340c
`
`[alpha]D = -96.5°
`
`Analysis calculated for C25H32C1N304S
`
`3 4
`
`5 6
`
`7 8
`
`9 Requires: c59.34 H6.37 N8.3o
`
`Found:
`
`059.51 H6.43 N8.24
`
`deltaH (250MHz, D6-DMSO) 8.85 (1H, s, NEOH), 8.37 (1H,
`d, J = 8.5Hz, CONE), 7.90 (13, m, CONgMe), 7.30 — 6.88
`
`(9H, m, aromatic H), 4-66 (1H, m, CECHZPh), 2.96 (1H,
`bd,
`J == 14Hz, CHCflzPh),
`2.76 (1H, bt,
`J == 13Hz,
`CHCflZPh) 2.60 (3H, d, J = 5Hz, NHCfl3), 2.55 — 2.40 (2H,
`m, CEZS), 2.16 (1H, m, cgc0), 2.01 ( 1H, d, J = 14Hz,
`CflCO), 1.37 (2H, m), 0.91 (1H, m, CEZCH(CH3)2), 0.81-
`(3H, d,
`J == 6Hz, CH(Cfl3)2),
`and 0.74 (3H,
`d,
`.J =
`
`6Hz,CH(CE3)2).
`
`(63.9MHz, D6-DMSO) 172.7, 171.6, 168.1, 139.2,
`deltac
`138.1, 130.3, 129.2, 127.9, 126.2, 125.9, 125.5, 125-0,
`54.1, 46.3, 45.8, 37.8, 32.0, 25.7,
`25.2,
`24.2,
`and
`
`21.7.
`
`10
`
`11
`
`12
`13
`
`14
`15
`16
`17
`18
`19
`
`20
`21
`
`22
`23
`24
`
`25
`
`26
`
`27
`
`28
`
`29
`
`30
`
`31
`
`32
`
`33
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0704
`
`page 0704
`
`

`

`WO 90/05719.
`
`7
`
`Pcr/GBs9/01399
`
`'54
`
`Example 18
`
`muO‘LfluhwMd
`
`11
`
`12
`
`13
`
`14
`
`15
`
`16
`
`17
`
`18
`
`19
`
`2o
`21
`
`22
`
`23
`
`24
`
`25
`
`26
`
`27
`
`28
`
`29
`
`[4—(N-Hydroxyamino)—2R—isobuty1—3s—(3é
`
`methylphenylthiomethyl) succinyl]-L—phenylalanine-N—
`methylamide
`
`I)
`
`
`
`Prepared by the method described in example 1g to give
`
`material with the following characteristics.
`
`Analysis calculated for C26H35N304S
`Requires: c64.3o H7.26 N8.65
`
`Found:
`
`€63.81 H7.21 N8.48
`
`.rdeltaH (250MHz, D6—DMSO) 8.83 (1H, s, NEQH), 8.35 (1H,
`d, J = 8.5H2, coma), 7.86 (1H, m, CONfiMe), 7.28 _ 6.77
`(9H, m, aromatic H), 4.6§ (1H, m, CECHZPh), 2;96 (1H,
`dd,
`J7= 14.4Hz, CHcg Ph), 2.80 (1H, bt, J = 13Hz,
`CHCfizPh) 2.59 (3H, d, J ; 5H2, NHcg3), 2.55 — 2.37 (2H,
`m, CEZS), 2.16 (2H, m, 2xcg00), 1.38 (2H, m), 0.91 (1H,
`m, CEZCH(CH3)2), 0.81 (3H, d, J
`6H2, CH(c§3)2), and
`0.74 (3H, d, J = 6H2, CH(cg3)2).
`
`3o_.
`31
`
`32
`
`33
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0705
`
`page 0705
`
`

`

`WO 90/05719
`
`PCT/GBS9/01399
`
`55
`
`Examgle 19
`
`[4—(N—Hydroxyamino)—2R—isobutyl—3S—(4—(N—acetyl)—
`
`aminophenylthiomethyl)succinyl]-L—phenylalanine—N—
`
`1
`
`2 3
`
`4
`
`5 methylamide.
`
`0
`
`Ph
`
`¢H
`“
`
`O
`
`H
`N
`
`NHMe
`
`
`
`CONHOH
`
`S
`
`ACNH
`
`A)
`
`[2R—isobutyl—38-(4—aminophenylthiomethyl)succinyl]—
`
`L—phenylalanine -N—methylamide.
`
`6
`
`7
`8
`
`9.
`
`10
`
`11
`12
`
`13
`14
`
`15
`
`16
`
`17
`
`18
`
`19 Prepared by the method described in example 1f to give
`
`20 material with the following characteristics.
`
`21
`
`22
`23
`24
`25
`
`deltaH (250MHz, D6—DMSO) 8.27 (1H, d, J = 8.5Hz, CONg),
`7.81
`(1H, m, CONflMe), 7.30 - 7.00 (5H, m, phenyl H),
`6.86 (2H,
`d,’ J '= 8.5Hz, aromatic H), 6.45 (2H,'d, J =
`8.5Hz, aromatic H), 5.25 (1H, bs, mg). 4.48 (1H, m,
`
`26 CgCHZPh), 2.91 (1H, dd, J = 14,4Hz, CHCflzPh), 2.88 (1H,
`
`27
`
`dd, J = 14,10Hz, CHCEZPh) 2.56 (3H, d, J = 5H2, NHCfl3),
`
`28
`2.43 — 2.24 (3H, m, C323 and chO), 2.03 (1H, d, J =
`29 ,10Hz' CECO), 1.41 (1H, t, J = 11Hz, CEZCH(CH3)2), 1.26
`30
`”H: mr CH2C§(CH3)2), 0.85 (1H; m, CH2CH(CH3)2), 0.81
`
`31
`
`(3H,
`
`d”
`
`J = 6H2, CH(Cfi3)2),
`
`and 0.74 (3H, d,
`
`J=5Hz,
`
`32 CH(CE3)2)-
`
`33
`
`‘
`
`AQUESTIVE EXHIBIT 1004 page 0706
`
`AQUESTIVE EXHIBIT 1004 page 0706
`
`

`

`' wo 90/05719 '
`
`'
`
`'
`
`,
`
`,
`
`56
`
`PCT/GBS9/01399
`
`[2R-isobfityl43S-(4-(N—acetyl)aminophenyl-thio—
`B)
`1
`'2 methyl)— succinyl]—Lphenyla1anine-N—methylamide.
`
`3,
`
`,
`
`4 The product from above (350mg, 0.74 mmol) was dissolved
`, 5
`in DCM (5 ml) cooled in an ice bath then triethylamine
`'6
`(75mg, 0.74 mmol), DMAP
`(91mg, 7.4 mmol) and finally
`7 acetic anhydride (83mg, 8.2 mmol) were added and the
`8
`solution stirred at RT for 90 minutes.
`The mixture Wasp
`
`9 partitioned between ethyl acetate and citric acid then
`10
`the organic layer washed with water and finally dried
`
`11 Over magnesium sulphate. Solvent removal gave the crude
`12 product as pale yellow crystals (160mg, 0.31 mmol,
`13
`742%).
`g
`
`14
`15
`16
`17
`18
`
`.
`7
`deltaH 7(250MHz, D6—DMSO). 9.94 (1H, s, COZH), 8.34'(1H,
`d, J =78.5Hz, CONE), 7.90 (1H, m, CONgMe), 7.46 (2H, d,
`J = 8.5Hz, aromatic H) 7.30 - 7.00 (5H, m, phenyl H),
`6.96 (2H, d, J a 8.5Hz, aromatic H), 4;57 (1H, m,
`
`chHZPh), 2.91 (1H,,dd, J = 14,4Hzr, CHcgzph), 2.88 (1H,
`19
`20 bt, J,= 13Hz; CHngPh), 2.58 (3H, d, J = 5H2, NHCfl ),
`21
`2.43 - 2.16 (3H, m, C525 and QECO), 2.10 (1H, d, J a
`22
`14Hz, chO), 17.354111,
`t,’ J = 1432, CHZCH(CH3)2), 1,25
`23
`(1H, m, CHZCg(CH3)2), 0.86 (1H, m, CEZCH(CH3)2), 0.81
`24
`(3H,
`.d,'J = 6Hz,fCH(C_I§3)2),
`and 0.74 (3H,rd, J =
`25'
`’6Hz,CH(cg3)2).
`'
`
`26
`27 ,C)
`28
`
`29
`
`,
`,
`7
`,
`,
`[4-(NrHydroxyamino)—2R—isobutyl-3S—(4-(N—acetyl)—
`aminophenylthiomethyl)succinylJ-L—phenylalanine-N_
`
`methylamide.
`
`,
`
`'
`
`,
`'
`,
`30
`31 Prepared by'the method described in example lg to give
`32 material with the following characteristics.
`
`33
`
`.
`
`'
`
`:
`
`a
`
`*
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0707
`
`page 0707
`
`

`

`WO 90/05719
`
`PCT/GB89/01399
`
`57
`
`m.p. 201 —202°c (dec.)
`
`[alphaJD = -7.50 (C=1.0, methanol)
`
`deltaH (250MHz, D6—DMSO) 9.90 (1H, s, NHQE), 8.02 (1H,
`s, NEOH), 8.30 (1H, d, J = 8.5Hz, CONE), 7.85 (1H, m,
`CONflMe), 7.45 (2H, d, J = 8.5Hz, aromatic H), 7.28 —
`
`6.94 (5H, m, phenyl H), 6.90 (2H, d,
`J = 8.5Hz,
`aromatic H), 4.66 (1H, m, CflCH2Ph), 2.90 (1H, dd, J =
`
`14,4Hz, CHcgzPh), 2.76 (11% bt,
`
`J = 13Hz, CHcgzph),
`
`2.50 (3H, d, J = 5H2, NHCfi3), 2.49 — 2.35 (2H, m,
`chS), 2.14 (1H, m, chO), 2.03 (4H,
`s + m, COCH3 and
`CfiCO), 1.35 (2H, m), 0.86 (1H, m, cgzcs(ca3)2), 0,81
`
`(3H, d, J-= 6H2, CH(c§3)2), and 0.74 (3H, d, J = 6H2,
`
`mHmWN'D-LQN"
`
`9
`
`10
`
`11
`12
`13
`
`14
`
`15 CH(333D)-
`16
`
`1 7 Examgle 20
`
`18
`
`19
`
`[4—(N—Hydroxyamino)—2R—isobutyl—3S—phenylsulfinyl—
`
`20 methylsuccinyl]—L—phenylalanine—N—methylamide.
`
`21
`
`22
`
`23
`
`24
`
`25
`
`26
`
`27
`
`28
`
`29
`
`30
`
`31
`
`32
`
`33
`
`
`
`'
`
`[4-(N—Hydroxyamino)—2R-isobutyl-38-phenylthiomethy1-
`
`succinyl]-L-phenylalanine—N-methylamide
`
`(250mg,
`
`0.53mmol) was dissolved in methanol
`
`(50 ml) and meta-
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0708
`
`page 0708
`
`

`

`WO 90/05719
`
`j
`
`7
`
`'
`
`Pcr/GBs9/01399
`
`58
`
`chloroperbenzoicr acid (100mg,
`
`0.58 mmol) was added.
`
`temperature ether was
`After stirring for 111 at.
`room,
`Solvent removal gave
`added and the mixture filtered.
`recrystallised fromr
`the crude. white solid. which was
`methanol / water then slurried in ether to remove final
`
`traces of meta-chlorobenzoic acid to give the desired
`
`material
`
`(70 mg, 0.014 mmol, 27%).
`
`fl
`
`m.p. 186 —1880c
`
`[alphaJD = -13.60 (c=0.5, methanol)
`
`Analysis calculated for C25H33N3OSS.O.SH20
`Requires:
`€60.46 H6.90 N8.46
`
`Found:r
`
`C60.58 H6.69 N8.29
`
`deltaH (250MHz, D6-DMSO, mixture of diastereomers) 9.04
`+ 8.93 (1H, 2x3, nng), 8.29 + 8.16 (1H, 2xd, J = 8.5
`Hz, Cong), 7.79 (1H, m, CONgMe), 7.90 — 7.40 (8H, m,
`aromatic H), 7.06 + 6.82 (2H,
`2xm, SO-Aromatic), 4.37
`(1H, m, CECHZPh), 2.93 _ 2.58 (3H, m, containing
`cncgzph), 2.52 (3H, m, NHQ§3), 2.49 + 2.37 (1H, 2xm),
`1.49 - 1.25 (2H, m, CHZCH(CH3)2 and cnzcg(CH3)2), 0.95
`(1H, m, CEZCH(CH3)2 ), 0.81 (3H, d, J
`6H2, CH(Q§3)2),
`and 0.74 (3H, d, J=6Hz, CH(Q§3)2).
`
`(63.9MHz, Dé—DMSO, mixture of diastereomers)
`deltaC
`172.2, 171.4, 171.3, 167.7,V144.5, 138.0, 137.9, 131.3,
`130.9, 129.6, 129.3, 129.1, 128.8, 128.3, 127.8, 126.5,
`126.2, 124.3, 123.6, 59.8, 58.1, 54.3, 54.0, 46.2,
`45.8, 41.6, 40.9, 37.6, 37.4, 25.6, 25.0, 24.3, 24.2,
`21.7, and 21.6.
`
`mummaawN—I
`
`Low—40mm\Immmwmaommqmmawm‘mom
`
`U
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0709
`
`page 0709
`
`

`

`WO 90/05719,
`
`'
`
`PCF/GBSQ/01399
`
`59
`
`Example 21
`
`[4—(N—Hydroxyamino)-2R—isobutyl—38—phenylsulfonyl—
`
`1
`
`2 3
`
`4 methylsuccinyl]—L—phenylalanine-N—methylamide.
`
`
`
`[4-(N-Hydroxyamino)—2R—isobutyl—3S—phenylthiomethyl—
`
`succinyl]-L-phenylalanine-N-methylamide (50mg,
`
`0.11mmol) was dissolved in methanol
`
`(12 ml) and meta-
`
`chloroperbenzoic acid (40mg, 0.23 mmol) was added.
`
`5 6 7 8 9
`
`10
`
`11
`
`12
`
`13
`
`14
`
`15
`
`15
`
`17 After stirring for 3h at
`
`room temperature ether was
`
`added and the mixture filtered.
`
`Solvent
`
`removal gave
`
`the crude white solid which was slurried in ether to
`
`remove final traces of meta—chlorobenzoic acid to give
`
`the desired material.
`
`18
`
`19
`
`20
`
`21
`
`22
`
`23 m.p. 228 — 231°C
`
`24
`25
`26
`
`[alpha]D = 16.80 (c=o.5, methanol)
`
`27 Analysis calculated for C25H33N3068.O.3H20
`
`28 Requires: C58.99 H6.65 N8.25
`
`29
`
`30
`
`31
`
`32
`
`33
`
`Found:
`
`€58.92 H6.51 N8.05
`
`deltaH (250MHz, D6—DMSO) 8.66 (1H, s, NEOH), 8.25 (1H,
`
`d, J = 8.5 Hz, CONE), 7.83 (1H, m, CONgMe), 7.75 — 7.50
`
`(5H, m, aromatic H), 7.30 7.05 (5H,
`
`nu
`
`aromatic H),
`
`AQUESTIVE EXHIBIT '1004 page 0710
`
`AQUESTIVE EXHIBIT 1004 page 0710
`
`

`

`WO 90/05719
`
`,
`
`7
`
`60
`
`PCI'/G389?01399
`
`mq‘mUw.ptuNA
`
`WMWLANNNJUNNNNNNA—‘A—‘J-‘A-J—‘-—"
`wN—Iowooxlmmth—Iommqmmth—‘om
`
`4.35 (1H, m, cgcn2ph); 2.86 (1H, dd, J = 14,5 Hz,
`CHCfiZPh), 2.75 (1H, dd, J = 14,10 Hz, CHcgzPh), 2.54
`(3H, d, J = 4.5 Hz, NHcg3), 2.54 (23, m), 1.30 (25, m,
`
`CEZCH(CH73)2 a’nd.CH2cg(CH3)2),
`
`0.86 (1H, m,
`
`fl
`
`chCH(CH3)2), 0.75 (3H, d, J = 6H2, CH(C§3)2), and 0.71
`
`(3H, d, J = 6Hz,cn(cg3)2).
`
`ExamEle 22
`
`[ 4-( N5-Enyir'd}{y'anliIIO')-2 R —:Ls olau.t§rl-—3 S —
`
`thiophenylsulphinylmethyl—succinyl]
`methylamide
`
`-L-phenylalanine—N—
`7
`
`
`
`[4—(N—Hydroxyamino)e2R-isobutyl-3S-thiophenylthio~
`methyl—succinYlJ-L?phenYlalanine-N—methylamide (50mg,
`0.11mmol) Was
`treated as described in example 21
`to
`yield the title compound (16mg, 0.03 mmol,
`29%) as a7
`mixture of «iiastereomer VWith.
`the following
`
`characteristics;
`
`m.p- 195 -197°c (dec.)
`
`Analysis calculated for C23H31N30582.0.5H20
`Requires: 054.96 H6.42 N8.36
`
`Found:
`
`7C54.91 H6.23 N8.23
`
`7 AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0711
`
`page 0711
`
`

`

`WO 90/05719
`
`PCT/GBS9/01399
`
`61
`
`1 2
`
`deltaH (250MHz, D6—DMSO, mixture of diastereomers) 9.04
`+ 8.96 (1H, 2X5, NEOH), 8.34 + 8.29 (1H, 2xd, J = 8.5
`3 Hz,
`CON_I_-I_), 8.02 + 7.98 (1H, 2xm, CONggMe), 7.81 (1H, bs,
`
`4
`
`thiophene—H), 7.42 (1H,
`
`s,
`
`thiophene-H), 7.25 — 7.15
`
`thiophene—H), 4.43 (1H,
`(5H, m, phenyl), 7.03 (1H, bs,
`5
`6 m, CfiCH2Ph), 3.0 - 2.6 (4H, m,
`containing CHCEZPh),
`7
`2.52 (7H, m, containing NHCE3), 2.05 (1H, m), 1.6 ~ 1.2
`
`8
`(2H, m, CflZCH(CH3)2 and CHZCE(CH3)2), 0.87 (1H, m,
`9 CEZCH(CH3)2), and 0.85 — 0.71 (6H, m, CH(C§3)2).
`1O
`
`11 W
`
`12
`
`13
`
`14
`
`I4-(N-Hydroxyamino)~2R-isobutyl—3S—
`
`thiophenylsulphonylmethyl-succinyl]
`
`-L—phenylalanine—N—
`
`15 methylamide.
`
`16
`
`17
`
`18
`
`19
`
`20
`
`21
`
`2:
`
`24
`25
`
`Z I Z 0
`
`221:
`
`CONHOH
`
`O
`
`
`
`U
`
`[4—(N-Hydroxyamino)-2R;isobutyl-3S-thiophenylthio—
`
`25 methyl-succinyl]—L-phenylalanine—N—methylamide (75mg,
`
`27
`
`0.16mmol) was
`
`treated as described in example
`
`22
`
`to
`
`28 yield the title compound (40mg, 0.08 mmol,
`
`49%) with
`
`the following characteristics:
`
`29
`
`30
`
`31 m.p. 215 — 216°C
`
`32
`
`33 Analysis calculated for C23H31N30682
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0712
`
`page 0712
`
`

`

`WO 90/05719
`
`7'
`
`.
`
`7
`
`'
`
`62
`
`- PCT/GBS9/01399
`
`71 Requires: c54.21 H6.13 N8.24
`
`2 Found:
`3
`
`054.07 H6.19 N8.04
`,
`,
`
`,
`
`4
`5
`5
`
`06—DMSC) 887 (1H, s, NgoH), 8.25 (1H,
`deltaH (250MHz,
`d,
`.T. = 8.5 Hz, CONE), 8.09 (1H, d,
`J .-. 4.7 Hz,
`thiophene-H), 7.83 (1H, m, CONEMe), 7.53 (1H, d, J = 3.
`
`:1
`
`7 Hz,
`
`thiophene H), 7.25 - 7.12 (6H, m, phenyl and
`
`thiophene—H), 4.36 (1H,rm, CECH2Ph), 3.38 ('1H, dd, J =
`8
`14,11 Hz, 8032), 2.87 (1H, dd, J = 14,5 Hz, CHcgzph),
`9
`10 '2.75 (1H, dd, J = 14,10 Hz, CHCgZPh), 2.70 — 2.36 (6H,
`11 m, containing NHCfl3), 1.20 (2H, m, CEZCH(CH3)2 and
`12 CHZQ§(CH3)2), 0.89 (1H,m, C§2CH(CH3)2 ), and 0.75 (6H,
`13 m, CH(C§3)2).
`14
`.
`
`,
`
`7
`
`7
`
`-
`
`15
`15
`17
`
`deltaC (63.9MHz,,06-DMSO) 172.0, 171.2, 166.5, 140.0,
`138.0, 135.4, 134.6, 129.0, 128.4, 128.2, 126.6, 54.3,
`45.6, 37.5, 25.6, 25.0, 24.2, and 21.7.
`'
`
`18 7
`
`19
`
`20
`
`21
`
`7
`
`§§2£2£2_;é
`
`,
`
`[4—(N-HydroxYamino)-2R—isobutyl—3S—phenylsulfonyl-
`
`22 methylsuccinyl]—L—phenylalanine—N-methylamide sodium
`
`23
`
`24
`
`25
`26
`
`27
`28
`
`29
`
`30
`
`31
`
`32
`33
`
`salt.
`
`7
`
`Ph
`
`&fl
`
`'
`
`o
`
`NHM;
`
`
`
`CONHONa
`
`,
`,
`[4—(N—Hydroxyamino)—2R-isobutyi—3S-phenylsulfonyl-
`
`1
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0713
`
`page 0713
`
`

`

`WO 90/05719
`
`PCT/GB89/01399
`
`63
`
`1 methylsuccinyl]-L—phenylalanine—N-methylamide (50mg,
`
`2
`
`0.1mmol) was dissolved in methanol
`
`(10ml)
`
`and sodium
`
`3 hydroxide solution (0.1M,
`
`1.0ml)
`
`added to give a
`
`homogeneous solution.
`
`The methanol was
`
`removed under
`
`reduced pressure then the residual
`
`aqueous
`
`solution
`
`freeze dried to give the title compound (40mg).
`
`deltaH (250MHz, D6-DMSO) 8.66 (1H, s, Ngon), 8.25 (1H,
`d, J = 8.5 Hz, CONE), 7.83 (1H, m, CONfiMe), 7.75 — 7.50
`
`4
`
`5
`
`6
`
`7 8
`
`9
`
`10
`
`(5H, m, aromatic H), 7.30 7.05 (5H, m, aromatic H),
`
`11
`4.36 (1H, m, CECHZPh), 2.86 (1H, dd,
`J = 14,5 Hz,
`12 CHCflzPh), 2.75 (1H, dd,
`J = 14,10 Hz, CHCEZPh), 2.54
`13
`(3H, d,
`J=4.5 Hz, NHCE3), 2.54 (2H, m), 1-30 (2H, m,
`
`m,
`(1H,
`0.86
`and CHZCE(CH3)2),
`14 CEZCH(CH3)2
`15
`chCH(CH3)2), 0.75 (3H, d, J = 6H2, CH(CE3)2), and 0.71
`16
`(3H, d, J = 6H2, CH(C§3)2).
`
`17
`
`18 W
`
`19
`
`20
`21
`
`22
`
`23
`
`24
`
`25
`
`26
`
`27
`
`28
`
`LONOKO
`
`but» N...‘
`
`DJ 00
`
`[4-(N—Hydroxyamino)—2R—isobutyl—BS—(4—(isobutyloxy—
`carbonylamino)phenyl)thiomethyl—succinyl]—L-pheny1—
`
`alanine-N—methylamide
`
`
`
`Z I
`
`4’
`
`a)
`
`[4—Hydroxy—2R—isobutyl-38-(4-aminophenyl)thio—
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0714
`
`page 0714
`
`

`

`’
`
`‘
`
`wo 90/05719
`
`7
`
`'
`
`PCT/GB89/01399
`
`,64
`
`1 methylsuccinyl]~Lephenylalanine—Nemethylamide was
`2 prepared by the method described in example 1i to give
`3
`a compound with the following characteristics.
`
`,
`,
`,
`_
`,4
`5 deltaH (250MHz, Ds-DMSO) 8.26 (1H, d, J7: 8.5 Hz,
`coma), 7.817 (1H, m, congMe), 7.27 - 7.15 (5H, m, phenyl
`
`H), 6.85 (2H, d, J = 8.5Hz, aromatic H), 6.46 (2H, d, J
`= 8-5Hz, aromatic H), 5.2 (1H, bs, c023), 4.48 (1H, m,’
`cgcaZ—Ph), 2.90 ('1H, dd, J ='13.5,4.3 Hz, CHcgzPh), 2.75
`9
`(1H, dd, J = 13.6, '10 Hz, CHCflzPh), 2.56 (3H, d, J =
`10
`.4.5 Hz, NHCE3).'2.S-Or—' 2.25 (3H, m), 2.03 (1H, d, J =
`1.1
`10 Hz),
`1.41 ,(TH, m, cgzcmcnyz),
`1.25 (1H, m,
`12
`13 CH2C§(7CH3)2), 0.86 (1H, m, chCH(CH3)2), 0.75 (3H, d, J
`14
`= 6H2, CH(c_1§3)2), and 0.71 (3H, d, J = ,6Hz,CH(C§3)2).
`15
`,
`16 lb) N,N4Dimethylglycine (100mg, 0.97 mmol) was stirred
`17
`in dry THF
`(50ml)
`and triethylamine (108mg,
`1.1mmol)
`18
`ahd isobutylchloroformate (146mg, 1.1mmol) were added.
`19 After 1h the product from example 26a (500mg, 1.1mmol)
`20 .was addedand the mixture stirred for a further 1h. The
`21
`reactithWas worked up by partitiofiing between citric
`22
`acid and ethyl acetate, drying the orgamic layer and
`23
`solvent rremoval
`to give the crude product
`(1g).
`24 Solution’ of, the crude solid inrethyl acetate then
`25 preCipitation withrether "resulted in white crystals of
`25
`the isobutlehloroformate derivative.
`,
`27
`7
`7,
`28
`[4—(N—Hydroxyamino)-2R—isobutyl—3S—(4-(isobutyloxy—
`c)
`29
`carbonylamino) phehyl)thiomethyl-succinylJ-L—phenyl—
`30' alanine-N-methylamide
`
`6 7 8
`
`'31
`
`,
`
`from example 26b was conVerted to the
`32 The product
`33
`hydroxamic acid as described in example 19.
`to give a
`compound with the following characteristics.
`
`7
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0715
`
`page 0715
`
`

`

`WO 90/05719
`
`PCT/GBS9/01399
`
`65
`
`1 m.p. 198 - 200°C
`
`[alpha]D = —8.50 (c=1, methanol)
`
`2 3
`
`4 5
`
`Analysis calculated for C30H42N4O6S
`6 Requires:
`€61.41 H7.22 N9.55
`
`Found:
`
`C62.04 H7.32 N9.67
`
`deltaH (250MHz, D6—DMSO) 9.60 (1H, s, NHOfl), 8.83 (1H,
`s, NgOH), 8.31 (1H, d, J = 8.5 Hz, CONE), 7.85 (1H, m,
`
`CONgMe), 7.36 — 7.25 (4H, m, aromatic H), 7.14 — 7.05
`
`(3H, m, aromatic H), 6.91 (2H, d, J = 8.5Hz, aromatic
`
`7
`
`8 9
`
`10
`
`11
`
`12
`
`13 H), 4.56 (1H, m, CECHZPh), 3.87 (2H, d,
`J = 7H2,
`14
`ochCH(CH3)2). 2.92 (1H, dd, J = 13.7,4.0 Hz, CHCflzPh),
`15
`2.76 (1H, dd, J = 13.6,10 Hz, CHCEZPh), 2.58 (3H, d, J
`16
`= 4.5 Hz, NHCE3), 2.50 — 2.34 (2H, m), 2.16 — 1.87 (3H,
`17 m),
`1.35 (2H,
`nu C§2CH(CH3)2 and CHéC§(CH3)2),
`0.93
`18
`(6H,
`d,
`J
`= 6.6H2, OCHZCH(C§3)2),
`0.87
`(1H,m,
`19
`C§2CH(CH3)2
`), 0.75 (3H, d,
`J = 6H2, CH(C§3)2),
`and
`20
`0.71 (3H, d, J = 6Hz, CH(C§3)2).
`21
`
`22
`
`23
`
`24
`25
`26
`27
`
`28
`
`29
`30
`
`31
`
`32
`
`33
`
`§§§22£§_2§
`
`.
`[4—(N—Hydroxyamino)—2R£isobuty1—3s—(4—(N—methy1_N_
`(tertbutoxycarbonyl)—glycylamino) phanyl)thiomethyl_
`succinyl]—Lphenylalanine-N-methylamide.
`
`Ph
`
`_-
`
`O
`
`H\
`.‘\\
`
`NHMe
`
`H
`N
`
`
`
`S
`
`O
`CONHOH -
`
`BOC\N/\fl/NH
`l
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0716
`
`page 0716
`
`

`

`\W()90/057]9
`
`,
`
`PCT/GB89/01399
`
`66
`
`ooqcxmpr—l
`
`wN-‘OLDCDQOXU'l-D-wN-JOkDmxlmlfl15b-LAJN-JO'LD
`
`wwwWNNNNNNNNNM—IA—IAAAJ—l—lg
`
`a)
`
`[4-Hydroxy-2R-isobutyl-3S-(4—(N—methyl—N—(tert—
`
`butoxycarbonyl)glycylamino) phenyl)thiomethyl-
`succinyl]—L-phenylalanine—N—methylamide was prepared as
`described in example 26b by substitution of N-BOC
`
`sarcosine for the acid component.
`
`)J‘
`
`deltaH (250MHz, D6-DMSQ) 9.97 (1H, s, cozg), 8.36 (1H,
`d, J'= 8.5 Hz, CONE), 7.91 (1H, m, CONHMe), 7.48 (2H,
`' a, J = 8.5Hz, aromatic H), 7.40 - 7.05 (5H, m, aromatic
`H), 6.97 (2H. d,rJ = 8.5Hz, aromatic H), 4.58 (1H, m,
`QHCHZPh), 3.95 (2H, d, J = 9Hz, NCHZCO), 2.92 (4H, m+d,
`CHQHZPh and Bocncg3), 2.76 (1H, dd, J = 13,10 Hz,
`CHQHZPH), 2.58 (3H, d, J = 4.5 Hz, NHCH3), 2.50 - 2.09
`(4H, m),
`1.46 - 1.33
`(11H,
`m -+ 2x5,
`(C§3)3C,
`CEZCH(CH3)72' and CHZCE(CH3)2),
`0.87 (1H, m,
`CHQCH(CH3)2'), 0.75 (3H, d, J = 6H2, CH(cH3)2), and
`0.71 (3H, d, J = 6H2, cH(cH3)2).
`
`[4—(N—Hydroxyamino)—2R—isobutyl—3S-(4-(N-methyl- N—
`b)
`(tertbutoxycarbonyl)-glycylamino)phenyl)- thiomethyl—
`succinylJ-Lphenylalanine—N—methylamide was prepared
`from the material produced in example 27a as described
`
`in example 1g.
`
`8.83
`cONHog),
`s,
`deltaH (250MHz, D6-DMSO)7 9.97 (1H,
`(1H, s, NHQH), 8.32 (1H, d, J = 8.5 Hz, CONE), 7.85
`(1H, m, CONHMe), 7.45 (2H, d; J = 8.5Hz, aromatic H),
`7.28 - 7.00 (5H, m, aromatic H), 6.97 (2H, d, J 4
`8.5Hz, aromatic H), 4.56 (1H, m, CECHZPh), 3.94 (2H, d,
`' J =79Hz, NCHZCO), 2.87 (4H, m+d, CHC§2Ph and BOCNcH3),
`2.76 (1H, m, chHZHh), 2.57 (3H, d, J = 4.5 Hz, NHQH3),
`2.25 ._ 1,91 '(2H, m), 1.42 -— 1.30 (11H,
`1n
`+ 2x5,
`
`d1
`qucH(CH3)2 ):,°~8° (3H.
`70.73 (3H, d, J=6Hz, CH(CH3)2).
`
`‘7 = 6H2. CH(CH3)2),
`
`and
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0717
`
`page 0717
`
`

`

`WO 90/05719
`
`PCF/G389/01399
`
`67
`
`Example 27
`
`Collagenase inhibition activity
`
`The potency
`
`of
`
`compounds of general formula I to act
`
`as inhibitors
`
`of
`
`collagenase (a. metalloproteas
`
`involved in tissue degradation) was determined by the
`
`procedure of Cawston and Barrett,
`
`(Anal- Biochem., 99,
`
`1 2
`
`3 4
`
`5 6
`
`7
`
`8
`
`9
`
`10
`
`340—345, 1979), hereby incorporated by reference,
`
`11 whereby a 1mM solution of the inhibitor being tested or
`
`12 dilutions thereof was
`
`incubated
`
`at 370 for 16 hours
`
`13 with collagen and collagenase (buffered with 25mM
`
`35 and
`0.05% Brij
`pH 7.5 containing 5mM CaClZ,
`14 Hepes,
`15
`0.02% NaN3).
`The collagen was acetylated 14C collagen
`16
`prepared by the method of Cawston and Murphy
`(Methods
`
`17
`
`18
`
`19
`
`20
`21
`
`in Enzymology, 80, 711, 1981), hereby incorporated by
`
`reference.
`
`The
`
`samples were centrifuged to sediment
`
`undigested collagen and an aliquot of
`
`the radioactive
`
`supernatant
`counter as
`
`removed for assay on
`a measure
`of hydrolysis.
`
`a scintillation
`The collagenase
`
`22 activity in the presence of
`
`1
`
`mM inhibitor,
`
`or
`
`a
`
`23 dilution thereof, was compared to activity in a control
`
`inhibitor and the results reported below as
`devoid of
`that
`inhibitor concentration effecting 50% inhibition
`
`of the collagenase (IC50)'
`
`Compound of Example No.
`
`1
`2
`2
`
`EESO
`
`20 nM
`8 nM
`73 nM
`(50% @
`1 mcM)
`
`24
`25
`
`26
`27
`
`28
`
`29
`30
`31
`32
`
`33
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0718
`
`page 0718
`
`

`

`wo 90/0571’9
`
`,
`
`Pcr/GBS9/01399
`
`68
`
`2
`
`3
`
`,Example 28
`
`7'
`
`'
`
`4 Stromelysin inhibition activity
`
`a
`'
`_
`'
`5
`The potency of compounds of general formula I'to act as
`6
`7 rinhibitors of stromelysin was determined using the
`8' procedure of Cawston g: g; (Biochem. g., 195,
`159—165
`9
`1981), hereby incorporated by reference, whereby a 1mm
`10
`solution. ofr
`the inhibitor being tested; or dilutions
`11
`”thereof: was
`incubated at. 37°C ‘for
`16 hours with
`127
`stromelysin and 14C acetylate casein (buffered with
`13
`ZSmM Hepes, pH 7.5 containing SmM CaClz, 0.05% Brij 35
`14
`and O;02%77NaN3Q
`The
`rcasein was
`14'C acetylated
`15
`according to the method described in Cawston gt a;
`15 “(Biochem.lg., 195, 159-165, 1981), hereby incorporated
`17
`by reference.r The stromelysin activit§ in the presence
`187 of 1mM, or a dilution thereof, was composed to activity
`19
`in’a control devoid of
`inhibitor and the results
`20
`reported below as
`that
`inhibitor concentration
`21
`effecting 50% inhibition of the stromelysin (ICSO).
`227’
`7
`'
`23
`Compound of Example No.
`24
`1
`25
`2
`26
`,
`7
`_
`27 Examples of unit dosage compositions are as follows:
`
`a
`
`,
`
`2950
`10 nM
`20 nM
`
`F
`
`“
`
`s
`
`a
`
`iii
`
`28
`29
`
`30
`
`31
`32
`
`0.!
`
`‘L0
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0719
`
`page 0719
`
`

`

`WO 90/05719
`
`PCT/GBS9/01399
`
`69
`
`Example 29
`
`Capsules:
`
`Ingredients
`
`Per Capsule
`
`Capsules
`
`Per 10,000
`
`1.
`
`Active ingredient
`
`de. of Form. I
`
`40.0 mg
`
`2.
`
`3.
`
`Lactose
`
`Magnesium
`
`stearate
`
`150.0 mg
`
`7
`
`4.0 mg
`
`194.0 mg
`
`400 g
`
`1500 g
`
`40 g
`
`1940 g
`
`Procedure for capsules:
`
`Step 1.
`
`Blend ingredients No.
`
`1 and No.
`
`2 in a
`
`suitable blender.
`
`Step 2.
`
`Pass blend from Step 1
`
`through a No. 30 mesh
`
`(0.59 mm) screen.
`
`Step 3.
`
`Place screened blend from Step 2 in a
`
`3 and
`suitable blender with ingredient No.
`blend until the mixture is lubricated.
`
`Step 4.
`
`Fill into No.
`
`1 hard gelatin capsule shells
`
`on a capsule machine.
`
`(DflmthJM-J
`
`wwwwNNNNNNNNNN—‘A—‘AA—‘dgdgwN—‘OLDCOQOU‘IfiUJNAOKOmflmthN-‘OKD
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0720
`
`page 0720
`
`

`

`W0 90/05719
`
`PCT/GB89/01399
`
`70
`
`Loooqmwthr-J
`
`uI»0:un)Nn)mn:NhJNn:N+4H+4H+4H+4H+4Hwh314o\om~JmU1pL»N+4otom.4m01p0:N+4o
`
`Ersnsls.§9
`
`Tabletsé
`
`'
`
`,
`
`w;
`
`Per 10,000
`
`Ingredients
`
`Per Tablet
`
`Tablets
`
`1.
`
`Active ingredient
`09d. of Form. I
`Corn Starch
`2.
`3- Alginic acid
`4.
`Sodium alginate
`5.
`’Magnesium
`
`40.0 mg
`720.0 mg
`7 20.0 mg
`20.0 mg
`
`stearate'
`7‘
`
`1.3 mg
`101.3 mg
`
`'
`
`400 g
`200 g
`200 g
`200 g
`
`r
`
`'
`
`,
`
`13 g
`1013 g
`
`Procedure for tablets:
`
`Step 1.
`7
`
`.Blend ingredients No. 1, No. 2, No.
`4 in a suitable mixer/blender.
`
`3 and No.
`
`Step 2.
`
`Add sufficient water portionwise to the blend
`
`7 from Step 1 with careful mixing after each
`
`addition.
`
`'Such additions of water and mixing
`
`7 until the mass is of a consistency to permit
`its conversion to wet granules.
`The wet mass is.converted to granules by
`
`Step 3.
`
`' passing it through an oscillating granulator
`7 uSing a No. 8 mesh (2.38) Screen.
`7
`
`Step 4.
`
`The wet granules are then dried in an oven at
`
`V140°F (60°C) until dry.
`
`Step 5.
`
`The dry granules are lubricated with
`
`'
`Step 6.
`
`ingredient No. 5.
`The lubricated granules are compressed on a
`suitable tablet press.
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0721
`
`page 0721
`
`

`

`WO 90/05719
`
`PCT/GB89/01399
`
`71
`
`Example 31
`
`Intramuscular Injection:
`
`Ingredient
`Compound of Formula I
`
`1.
`
`Per ml.
`
`Per liter
`
`Active ingredient
`
`10.0 mg
`
`10 g
`
`2.
`
`Istonic buffer
`
`solution pH 4.0.
`
`q.s.
`
`q.s.
`
`Procedure:
`
`Step 1.
`
`Dissolve the active ingredient in the buffer
`
`solution.
`
`step 2. Aseptically filter the solution from Step 1.
`
`Step 3.
`
`The sterile solution is now aseptically
`
`filled into sterile ampoules.
`
`Step 4.
`
`The ampoules are sealed under aspetic
`conditions.
`
`Example 32
`
`Suppositories:
`
`1.
`
`2
`
`3
`
`Ingredients
`Compound of Form.
`
`I
`
`Per Supp.
`
`Per
`
`1‘000 Supp
`I
`
`Active ingredient
`
`40.0 mg
`
`40 g
`
`Polyethylene Glycol
`
`'
`
`1000
`
`1350.0 mg
`
`1,350 g
`
`Polyethylene Glycol
`
`4000
`
`450.0 mg
`
`1840.0 mg
`
`450 g
`
`1,840 g
`
`7
`
`1
`
`2 3
`
`4
`5
`
`6
`
`7
`
`8
`
`9
`
`10
`
`11
`
`12
`
`13
`
`14
`
`15
`
`16
`17
`
`18
`
`19
`
`20
`
`21
`
`22
`
`23
`24
`
`25
`
`26
`
`27
`
`28
`
`29
`
`30
`31
`32
`
`33
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0722
`
`page 0722
`
`

`

`wo 90/05719 7
`
`,7
`
`7
`
`PCT/GB89/01399
`
`72
`
`kDOZJ'dmLJI-thh-I
`
`wb)uo)Mn)NhJ‘Nh)Nh)N51Hp:Hpup;H‘paH+4HL»N+40L0m~JmLnoL»Mr4o\om~lmLnpu:m+4o
`
`Procedurei
`Step 1. VMelt ingredient No.
`
`_
`
`stir until uniform.
`
`2 and No.
`
`3 together and
`
`”I!
`
`Step 2.
`
`step'3;
`
`7
`
`DisSolve ingredient No. l in the molten mass
`from Step 1 and stir until uniform.’
`' Pour the molten mass from Step 2 into
`rsuppository moulds and chill.
`
`Step 4.’ Remove the suppositories from moulds and
`
`wrap.
`
`Exam le 33
`
`Eye'ointment
`
`An appropriate amount of a compound of general formula
`7 I,is formulated into an eye ointment base having the
`following compoSition:r
`
`Liquid paraffin
`
`,
`
`' Wool fat
`
`Yellow soft paraffin
`
`10%
`
`10%
`
`80%
`
`Example 34
`
`Topical skin ointment,
`
`An appropriate amount of a compound of general formula
`
`{I
`
`is formulated into a
`
`topical skin ointment base
`
`having the following composition:
`
`Emulsifying wax
`
`White soft paraffin
`
`,Liquid paraffin?
`
`o\°
`
`30
`
`50%
`
`o\°
`
`“I:
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0723
`
`page 0723
`
`

`

`WO 90/05719
`
`PCT/GB89/01399
`
`73
`
`CLAIMS
`
`1.
`
`A compound of general formula I:
`
`o
`
`R3
`
`R4
`
`N
`
`551
`R2
`I‘LH o
`T
`CONHOH
`RISOn'
`
`‘125
`
`(I)
`
`l
`
`2 3
`
`4 5
`
`7
`
`6
`8
`9,
`10
`
`11 wherein:
`
`12
`
`13
`14
`15
`16
`17
`18
`
`19
`
`MN Ho
`
`22
`23
`
`24
`
`25
`
`26
`
`27
`
`28
`
`29
`
`30
`
`31
`32
`33
`
`R1
`
`thiophenyl,
`phenyl,
`represents a C1—C6 alkyl,
`substituted phenyl, phenyl(Cl-C6)alkyl,
`heterocyclyl,
`(Cl-C6)a1kylcarbony1 or phenacyl or
`substituted phenacyl group: or when 11 = O, R1
`represents SRX, wherein RX represents a group:
`
`4
`
`U1
`
`m
`
`"'\ VZ/
`O
`
`CE'2'.
`CONHOH
`
`w
`
`A
`1
`
`R2
`
`represents a hydrogen atom or a Cl—C6 alkyl, Cl—C6
`
`a l k e n y l ,
`
`p h e n y l ( C 1 - C 6 ) a 1 k y l ,
`
`cycloalkyl(Cl-C6)alkyl or cycloalkenyl(Cl—C6)alkyl
`
`group;
`
`R3
`
`represents an amino acid side chein or a Cl-C6
`alkyl,
`benzy1,
`(C1—C6 alkoxy)benzyl or
`benzyloxy(Cl'—C6 alkyl) or benzyloxy benzyl group;
`
`AQUESTIVE EXHIBIT 1004 page 0724
`
`AQUESTIVE EXHIBIT 1004 page 0724
`
`

`

`WO 90/05719
`
`PCT/GB89/01399
`
`74
`
`‘kflmxlmU'lfi-WNH
`MH+4HFJFJH+4H+4HOLDOJQO‘UIWDUJNHO
`UNF-‘OkDODsImUIswal-J
`UHUUN‘NNNNNNNN
`
`represents. a hydrogen atom or a C1-C6 alkyl group;
`
`R5
`
`represents a hydrogen atom or a methyl group;
`
`In
`
`n
`
`A
`
`is an integer having the value 0, l or 2; and
`
`represents a C1-C6 hydrocarbon chain, optionaly
`substituted with one or more C1-C6 alkyl, phenyl
`or substituted phenyl groups;
`
`or a salt thereof.
`
`2.
`'A compound as claimed in Claim 1,
`in which the
`chiral. centre, adjacent
`the Substituent, R3 has
`S
`
`stereochemistry.
`
`A compound as claimed in Claim 1 or 2, wherein the
`3.
`chiral centre adjacent
`the substituent R2 has R
`
`stereochemistry.
`
`in which
`2 or 3,
`A compound as claimed in Claim 1,
`74.
`R1 represents a hydrogen atom or a Cl—C4 alkyl, phenyl,
`thiophenyl, benzyl, acetyl or phenacyl group.
`
`A compound as claimed in any one of Claims 1 to 4,
`5.
`wherein R2 represents a C3-C6Valkyl group.
`
`A_compound as claimed in any one of Claims 1 to 5,
`6.
`wlieax'e ir1 R3
`reap:reas erlt s
`a
`b eriz y l
`o r
`
`4—(Cl—C6)alkoxyphenylmethyl or benzyloxybenzyl group.
`
`A compound as claimed in any one of Claims 1 to 6,
`7.7
`wherein.R4 represents a C1-C4 alkyl groupi
`
`AQUESTIVE EXHIBIT 1004 page 0725
`
`AQUESTIVE EXHIBIT 1004
`
`page 0725
`
`

`

`‘P
`
`WO 90/05719
`
`PCT/GB89/01399
`
`75
`
`lomxlmUI-DLJNH
`
`utouL»Ntomh)MNDJMm)luHr4Hpa};H+4HHL»N+4o\om~4mLn»o:NF4otom~JmLnbLuN+4S
`
`8.
`A compound as Claimed in any one of Claims 1 to 7,
`wherein R5 represents a hydrogen atom.
`
`[4-(N-Hydroxyamino)-2R—isobutyl-BS-(phenylthio-
`9.
`methyl)-succinyl]—L—phenylalanine-N—methylamide,
`
`[4—(N—Hydroxyamino)-2R-isobutyl—BS-(thiophenylthio—
`methyl) succinyl]~L-phenylalanine-N—methylamide,
`
`[4—(N-Hydroxyamino)-2R-isobutyl—3S-(benzylthiomethyl)
`succinyl]-L-phenyla1anine-N—methylamide,
`
`[4-(N-Hydroxyamino)-2R-isobutyl—38-(acetylthiomethyl)
`succinyl]—L—phenylalanine-N—methylamide or
`
`[4-(N—Hydroxyamino)-2R-isobutyl-BS—(thiolmethyl)
`
`succinyl]-L-phenylalanine-N-methylamide
`
`[4—(N-Hydroxyamino)-2R—isobutyl-3S-(pivaloylthiomethyl)
`succinyl]-L-phenylalanine—N-methylamide
`
`[4—(N—Hydroxyamino)-2R—isobutyl—3S—(phenylthiomethyl)
`
`succinyl]-L—phenylalanine-N—methylamide sodium salt
`
`{4—(N-Hydroxyamino)—2R—isobutyl-38-(4-methoxyphenyl-
`thiomethyl)succinyl]—L-phenylalanine—N4methylamide
`
`[4-(N-Hydroxyamino)-2R-i§6butyl-BS-(4-hydroxypheny1—
`thiomethyl)succinyl]-L—phenylalanine—N—methylamide
`
`[4—(N-Hydroxyamino)-2R-isobutyl—38-(2fthiophenethio-
`methyl)succinyl]-L-phenylalanine-N-methylamide sodium
`salt
`
`AQUESTIVE EXHIBIT 1004
`
`AQUESTIVE EXHIBIT 1004 page 0726
`
`page 0726
`
`

`

`W0 90/05719
`
`I
`
`PCT/GB89/01399
`
`76
`
`[4—(N-Hydroxyamino)-2R—isobutyl-BS—(4—methoxyphenyl-
`
`thiomethyl)succinyl]-L—phenylalanine-N-methylamide
`sodium salt
`'7
`
`(“‘9
`
`[4?(N-Hydroxyamino)-2Reisobuty1-3S-(4-tertbutylphenyl-
`thiomethYl)succinyl]-Lephenylalanine-N—methylamide
`
`[4-(N—Hydroxyamino)—2R-isobutylé3s-(2,4-dimethylphenyl-
`thiomethyl) succinyl]-L-phenylalanine-N-methylamide
`
`r
`
`bis-S,S'e([4(N-Hydroxyamino—ZR-isobutyl-3S-(thiomethyl)
`
`succinyl]eL—phenylalanine-N—methylamide} disulphide
`
`'[4-(N-Hydroxyamiho)~2RriSobuty1—3S—(3-bromophenylthio—r
`methyl) succinyl]-L-phenylalanine-N-methylamide
`
`[4-(N—Hydroxyamino)-2R-isobutyl-3Se(3-chlorophenylthio-
`methyl) succinyl]-L—phenylalanine—N—methylamide
`
`r[4—(N-HYdroXyaminQ)-2R-isobutyl—3S-(3-methylphenylthio-
`methyl) succinyl]-L-phenylalanine-Némethylamide
`
`[4é(N4Hydroxyamino)-2R-isobutyl-3S-(4—(N—acetyl)—amino—
`
`phenylthiomethyl)succinyl]-L-phenylalanineéu-methyl

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