AU2003221318B2 - Synthesis of CC-1065/duocarmycin analogs - Google Patents

Description

AUSTRALIA

PATENTS ACT 1990 DIVISIONAL APPLICATION NAME OF APPLICANT: The Scripps Research Institute ADDRESS FOR SERVICE: DAVIES COLLISON CAVE Patent Attorneys 1 Little Collins Street Melbourne, 3000.

INVENTION TITLE: "Synthesis of CC-1065/duocarmycin analogs" The following statement is a full description of this invention, including the best method of performing it known to us: Q:\OPER\GC\12285950DIV 204.DOC 24/7/03 SYNTHESIS OF CC-1065/DUOCARMYCIN

ANALOGS

Description Technical Field: The present invention relates to a method for the synthesis of the dihydroindole C-ring found in CC-1065 duocarmycin analogs. More particularly, the invention comprises the 5-exo-trig radical cyclization of an aryl halide onto a tethered vinyl chloride forming the dihydroindole

C-

ring with chlorine installed as a suitable leaving group for subsequent cyclopropane spirocyclization. The versatility of this approach is examined in the context of six CC-1065 duocarmycin analogs previously synthesized in this laboratory.

Background: CC-1065 Chidester et al. J. Am. Chem. Soc. 1981, 1-3, 7629) and the duocarmycins 2 (Ichimura et al. J. Antibiot.

1990, 43, 1037) and 3 (Takahashi et al. J. Antibiot. 1988, 41, 1915; Yasuzawa et al. Chem. Pharm. Bull. 1995, 43, 378) are the parent members of a potent class of antitumor antibiotics that derive their biological properties through reversible, sequence selective alkylation of DNA (For a review of mechanistic aspects see: Boger, et al. Angew. Chem., Int. Ed.

Engl. 1996, 35, 230) Since their disclosure, synthetic efforts have focused on the natural products as well as a great number of rationally designed analogs (For a review of synthetic efforts see: Boger et al. Chem. Rev., 1997, 97, 787). These analogs have served define the fundamental principles underlying the relationships between structure, chemical reactivity and biological properties within this family, and have advanced the understanding of the origin of sequence selectivity and the catalysis of the DNA alkylation reaction by 1-3 (Boger et al.

J. Am. Chem. Soc. 1997, 119, 4977; Boger et al. J. Am. Chem.

11-08-'06 10:42 FROM- T-983 P006/025 F-815 POPEMpELPDBnupw nlitj pa mma imsw Va S-2- SSoc. 1997, 119, 4987; Boger et al. Biorg. Med. Chem. 1997, 263; Warpehoski et al. J. Am. Chem. Soc. 1994, 116, 7573; Warpehoski et al. J. Am. Chem. Soc. 1995, 117, 2951).

Common synthetic routes to many of the duocarmycin and 00 5 CC-1065 analogs incorporate the same transformation via a OC four step procedure highlighted by an in-situ trap of a C primary radical with TEMPO (TEMPO 2,2,6,6-tetramethyl-l- C(c piperidinyloxy free radical) followed by its reductive o removal and conversion to the chloride as depicted for the

C

10 synthesis of CBI (Boger et al. J. Org. Chem. 1995, 60, 1271) as illustrated in Figure 4.

It would be beneficial to have a more direct and higher yielding transformation to obtain the dihydroindole C-ring found in CC-1065 duocarmycin analogs. What is needed, therefore, is an efficient and general method for the synthesis of the dihydroindole C-ring round in CC-1065 duocarmycin analogs with less steps than the standard four step TEMPO procedure as described above.

Summary of the Invention: One aspect of the invention is a method for the synthesis of a dihydroindole C-ring of a CC-1065 duocarmycin analog wherein the method comprises the steps of: allylating an aryl halide with 1,3-dichloropropene in the presence of a catalytic amount of n-tetrabutylammonium iodide to form a vinyl chloride; then cyclizing the vinyl chloride conditions using tribuytyl tin hydride, catalytic AIBN and toluene as the solvent for forming the dihydroindole C-ring of the CC-1065 duocarmycin analog.

Another aspect of the invention is directed to the following compounds: COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:42 FROM- MCF0RSWOBpw?2US3IZDI hp.; IMM T-983 P007/025 F-815 -3-

N

O AN

BOG

HN I,

BOG

0 Meo

NC

OMe

CO

2 Me N Bn SNEr Bn HBC RCNB no NHBOC

CN

OMOM N C M eN 4 O 8 e B OGN N O~e 1*01

N

1 183n

N

Soc HN B CI

BOG

OMOM C1 MO N Me N O0k

SOC

COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08l1 11-08-'06 10:42 FROM- T-983 P008/025 F-815 IN -4- COMOM C1 b) MeOl OMe

BOO

Description of Fiscurs: 00 SFigure 1 shows CC-1065 and the duocarmycins and The compounds are parent members of a potent class of Ci &antitumor antibiotics.

Figure 2 shows CBI CPyI desmethyl-CPI iso-CBI and the mitomycin-hybrid which are compounds of interest in this application.

Figure 3 shows the novel intramolecular aryl radical cyclization onto a tethered vinyl chloride to install the dihydroindole C ring with chlorine installed as a suitable leaving group for subsequent cyclopropane spirocyclization.

Figure 4 shows the standard four step procedure highlighted by an in-situ trap of a primary radical with TEMPO (TEMPO 2,2,6,6-tetramethyl--piperidinyloxy free radical) followed by its reductive removal and conversion to the chloride as depicted for the synthesis of CBI.

Figure 5 illustrates the transformation from compound to 12a and the subsequent formation of a cyclopropyl ring.

Figure 6 shows a table which illustrates the results of the two-step synthesis of 3-chloro-methylindolines with the following conditions: a NaH, 1, 3 -dichloropropene, DMF, 25 °C; b NaH, 1,3-dichloopropene, nBu,NI, DMF, 25 c AIBN (cat.) Bu SnH, benzene, 60-75 C; d AIBN BuSnH, toluene, wherein each compound uses the same transformation as shown in Figure 6.

COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:43 FROM- T-983 P009/025 F-815 N D Detailed Description of the Inventn The invention is directed to a two-step transformation Sdirected to the synthesis of 6 CC-io65/duocarmycin analogs Susing a novel intramolecular aryl radical cyclization onto a vinyl chloride to form the dihydroindole C-ring found in 6 CC-1065/duocarmycin analogs. This transformation represents a OO potential two-step improvement to the synthetic route to many 0 other analogs, which most recently incorporated the same transformation via a four step procedure highlighted by an in- 10 situ trap of a primary radical with TEMPO (TEMPO 2,2,6,6- Stetramethyl-1-piperidinyloxy free radical) followed by its 0 reductive removal and conversion to the chloride as depicted for the synthesis of CBI (Boger et al. Org. Chem. 1995, 1271) as illustrated in Figure 4.

Patel et al. describes the synthesis of an analog named Oxa-duocarmycin SA which utilizes a novel intramolecular aryl radical cyclization onto a tethered vinyl chloride to install the dihydroindole C ring with chlorine installed as a suitable leaving group for subsequent cyclopropane spirocyclization as described in Figure 3.

Application of this improved two-step transformation to the synthetic routes reported for a number of the analogs synthesized in this laboratory would serve to establish the versatility of this approach to the synthesis of CC-1065 and duocarmycin analogs.

With this goal in mind, the C-ring construction for CBI (4; Boger et al. J. Org. Chem. 1995, 60, 1271, CCBI Boger et al.

J. Org. Chem. 1996, 61, 4894), CPyI desmethyl-CPI iso- CBI and the mitomycin-hybrid was investigated. The appropriately functionalized aryl halides (10a-g), which were obtained either through direct electrophilic halogenation (entries 1-5) or directed ortho metallation (entries 6 and 7) and halide quench, were allylated with 1,3 dichloropropene to COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:43 FROM- T-983 P010/025 F-815 P \ORWEA \Sposto200321 2. tW4oc-IId~tviJ \o Scomplete the radical cyclization precursors (lla-g) in high <yields. Treatment with Bu 3 SnH and a catalytic amount of AIBN (AIBN 2,2'-azobisisobutyronitrile) with heating in benzene or toluene very cleanly effected 5-exo-trig radical cyclization to form the 3-chloromethyl indoline C-ring present in each of the analogs (12a-g) as illustrated in Figure 6.

Cn e- This two-step transformation works well with benzene, C, naphthalene, indole and quinoline derivatives, aryl iodides as C, well as aryl bromides, with little to no deterioration in the O 10 consistently high yields for both steps. Brief optimization efforts revealed that higher yields may sometimes be obtained with addition of n-Bu 4 NI to the allylation reaction, as well as substitution of toluene and higher reaction temperature for benzene. It was observed, as also noted by Patel et al. that deoxygenation of the solvent prior to radical cyclization may enhance both the rate and yield of the reaction.

In summary, this novel intramolecular aryl radical cyclization onto a vinyl chloride, as introduced by Patel, was successfully applied to the C-ring synthesis of 6 CC- 1065/duocarmycin analogs. This application has effectively shortened the synthesis of each of these analogs by two steps.

Clearly the versatility of this approach, combined with the high conversions for both steps, assure its use in future rational analog design in the CC-1065/duocarmycin family of antitumor antibiotics.

While a preferred form of the invention has been shown in the drawings and described, since variations in the preferred form will be apparent to those skilled in the art, the invention should not be construed as limited to the specific form shown and described, but instead is as set forth in the following claims.

EXPERIMENTAL PROTOCALS feneral 1 H and "C nmr spectra were recorded either on a Bruker AM-250, a Bruker AMX-400 or a Bruker AMX-500 COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 -7spectrometer. Residual protic solvent CHC1, (5n= 7.26 ppm, 77.0), d.-methanol 3.30 ppm, 5c= 49.0) and D,O 4.80 ppm, 5c (of CHCN) 1.7 ppm) or TMS (5 0.00 ppm) were used as internal reference. Coupling constants were measured in Hertz HRMS were recorded using FAB method in a mnitrobenzylalcohol (NBA) matrix doped with Nal or CsI. Infrared spectra were recorded on a Perkin-Elmer FTIR 1620 spectrometer. Enantiomeric excess was determined by HPLC using a Daicel Chemical Industries CHIRALPAK AD column. Optical rotations were measured with an Optical Activity AA-1000 polarimeter. Melting points were taken on a Thomas Hoover capillary melting point apparatus and are uncorrected. Column chromatography was performed on Merck Kieselgel 60 (230-400 mesh). Analytical thin layer chromatography was performed using pre-coated glass-backed plates (Merck Kieselgel

F

2 54 and visualized by cerium molybdophosphate or ninhydrin. Diethyl ether, tetrahydrofuran (THF) and toluene (PhCH 3 were distilled from sodium-benzophenone ketyl, dichloromethane (DCM) and acetonitrile from calcium hydride. Other solvents and reagents were purified by standard procedures if necessary.

General Experimental Procedure for individual synthesis of 12a-g as shown in Figure 6: A solution of the aryl iodide (one of 10a-g as shown in Figure 6 obtained from the sources or conditions as described herein; aryl iodide is obtained from the following sources:) in anhydrous DMF (0.1M) at 0 oC was treated with NaH (2.0 equiv.) in small portions. The resulting suspension was stirred 15 min and treated with neat 1,3-dichloropropene (5.0 equiv) in a slow dropwise manner, followed by catalytic Bu,NI (0.1 equiv.; n-tetrabutylammonium iodide The reaction mixture was warmed to 25 OC and stirred for 12 h. The reaction mixture was quenched with the addition of 5% aqueous NaHC03, and the aqueous layer was extracted with EtOAc. The combined organic extracts were washed with H 2 0, dried (Na 2 SO4) and concentrated under reduced pressure. The crude was purified by flash column chromatography. A solution of one of lla-g in anhydrous benzene (0.1M; alternatively substitution of toluene and higher reaction temperature can optimize yield, due to higher temperatures) was treated with Bu 3 SnH (1.05 equiv.) and catalytic AIBN (0.1 equiv.) and deoxygenated with a stream of dry N 2 gas. The solution was heated to 80 OC for 2 h and concentrated in vacuo. The crude was purified by flash column chromatography to form one of compounds 12a-g.

Synthesis of cyclopropane via spirocyclization. The chlorine group of the dihydroindole C-ring is installed as a suitable leaving group for cyclopropane spirocyclization.

Methodologies for subsequent spirocyclization and further aklyation of the resultant cyclopropane C-ring system to the DNA portion of CC-1065 and the duocarmycins is well known in the art. A representative spirocyclization is accomplished via treatment of 12(a-g) with NaH (3 equiv, THF, 0 OC, 30 min) to provide as shown in Figure Similarly, acid-catalyzed deprotection of 12(a-g) (3N HC1-EtOAc, 25 OC, min) followed by spirocyclization of the crude indoline hydrochloride salt upon exposure to 5% aqueous NaHCO3-THF (1:1, OC, 1.5 h, 93%) can also provide as shown in Figure 2).

Synthesis of compounds 4-9 as shown in Figure 2 A solution of 12(a-g; one of the compounds in Figure 6; obtained from the sources or conditions as described herein) (1.5 mg, 4.1 Mmol) in tetrahydrofuran-dimethylformamide (3:1, 200 YL) at 0 OC under N 2 was treated with suspension of NaH mg, 60% in an oil dispersion, 12 4mol, 3 equiv). The reaction mixture was allowed to stir at 0 oC and for 30 min before the addition of pH 7 phosphate buffer (0.2 M, 250 /,zL) and 2 mL of tetrahydrofuran. The organic layer was dried (NaSO,) and concentrated in vacuo. Chromatography (SiO 2 20-30% Ethyl ace tate-hexane gradient elution) afforded (4-9; as shown in Figure Alternative Spyrocyclization. 12 (a-g; one of the compounds) (5 mg, 1.37 ;.mol) was treated with anhydrous 3N HC1 -Ethyl acetate (0.4 mL) at 24 OC for 20 min.

The solvent was removed in vacuo to afford the crude, unstable amine hydrochloride. This residue was treated with 5% aqueous NaHCO 3 (0.4 mL) and tetrahydrofuran (0.4 ML) at 24 0 C under

N.,

and the two phase mixture was stirred tor 1.5 h (24 OC) The reaction mixture was extracted with Ethylacetate (3 x 2 mL) and the combined extracts were washed with H.0 (2 dried (Na 2

SO

4 and concentrated in vacuo. Chromatography (SiO 2 CHOH-CHCl,) afforded as shown in Figure 2) OR

OMOM

H3C n-BuLi, TMEDA

H

3 CO Cu(N0 3 2

H

3 C -25 OC

H

3 C0* 8 H 3 C) R ICH 2

CH

2 CI, 74%

OCH

3

OCH

3 NaH MOMCI RJ~,l MO Al-Hg, 8 5 %Z'a 2 'PR N0 2 2 0, R- NHBOC OMOM OMOM

CI

H 3 C O I C Kl C H C o I NaH, 96%1

H

3 C NBOC

H

3 N OCH3 OC- OC 103 i 2,4-(Dimethoxy)-3- (methyl) -methoxymethyl phenyl ether (101).

A solution of 2 4 -(dimethoxy)-3-methylphenol (1.0 g, 5.95 mmol) in 60 mL of anhydrous DMF at 0 OC was treated with NaH (357 mg, 9.91 mmol) in several portions over 5 min. After min, Bu 4 NI (219 mg, 0.60 mmol) was added followed by the dropwise addition of C1CH 2

OCH

3 (0.68 mL, 8.91 mmol). The reaction mixture was stirred at 25 0 C for 36 h before the reaction was quenched by the slow addition of 30 mL of HO0.

The aqueous layer was extracted with EtOAc (3 30 mL) The organic layers were combined, washed with 10% aqueous NaHCO3 mL) and H,O (4 20 mL), dried (Na 2

SO

4 and concentrated under reduced pressure. Flash chromatography (SiO 2 3 10 cm, EtOAc/hexane) provided 101 (1.11 g, 88%) as a light yellow oil: 'H NMR (CDC1 3 250 MHz) 5 6.92 J 8.8 Hz, 1H), 6.51 J 8.8 Hz, 1H), 5.13 2H), 3.79 3H), 3.76 3H), 3.50 3H), 2.13 3H); 3 C NMR (CDC1 3 62.5 MHz) 5 153.5, 149.2, 144.3, 121.0, 114.4, 105.3, 96.0, 60.4, 56.0, 55.7, 8.9; IR (film) vmax 2937, 2833, 1595, 1487, 1440, 1420 cm-1; FABHRMS (NBA) m/z 212.1040 6 0 requires 212.1049).

2,4- (Dimethoxy) (methyl) (nitro) -methoxymethyl phenyl ether (102).

A solution of 101 (1.11 g, 5.21 mmol) in 18 mL freshly distilled Ac 2 O at 0°C was treated with Cu(N0 3 2 2.5 H 2 0 (2.41 g, 10.4 mmol) in several portions over 5 min. The reaction mixture was stirred for 2 h at 0 oC, then 1 h at 25 oC before the reaction was poured over H 2 0 (50 mL) and extracted with EtOAc (3 30 mL). The combined organic layers were washed with saturated aqueous NaC1 (50 mL),.dried (Na 2

SO

4 and concentrated under reduced pressure. The crude light yellow oil (1.18 g, 88%) was carried on to the next transformation:

'H

NMR (CDC13, 250 MHz) 6 7.54 1H), 5.18 2H), 3.87 (s, -11- 3H), 3.81 3H), 3.48 3H), 2.21 3H) "C NMR (CDC1 3 62.5 MHz) 5 153.0, 147.8, 145.8, 138.9, 128.2, 110.5, 95.3, 61.8, 60.5, 56.2, 9.5; IR (film) max 2942, 2829, 1522, 1481, 1344, 1280, 1246 cm- 1 FABHRMS (NBA) m/z 258.0977 (CHisNO 6 H* requires 258.0978).

(Amino) (dimethoxy) (methyl) -methoxymethyl phenyl ether (103).

A solution of 102 (1.18 g, 4.57 mmol) in 90 mL moist ether (8:2:1 Et 2 0:EtOH:H 2 0) was cooled to 0C, and treated with freshly prepared Al-Hg (1.23 g Al, 45.7 mmol) in small 1 1 cm pieces. The reaction mixture was stirred vigorously for h at 0 then 1 h at 25 The reaction mixture was then filtered through Celite, and the Celite was washed thoroughly with Et 2 O (5 20 mL). The solution was then washed with saturated aqueous NaCl (100 mL), dried (Na 2

SO

4 and concentrated under reduced pressure to afford 103 (0.88 g, as a crude brown oil, which was immediately carried on to the next step: IH NMR (CDC13, 250 MHz) 5 6.42 1H) 5.11 (s, 2H), 3.70 3H), 3.66 3H), 3.56 2H), 3.47 3H), 2.16 3H) "C NMR (CDC1 3 250 MHz) 5 147.0, 140.4, 140.3, 135.8, 125.4, 102.0, 95.4, 60.6, 59.4, 56.0, 9.34; IR (film) vm,, 3446, 3359, 2935, 2826, 1617, 1492, 1358 cm-1; ESIMS m/z 228 H* requires 228).

(tert-Butyloxycarbonyl)amino] (dimethoxy) (methoxymethoxy) -3-methylbenzene (104).

A solution of crude 102 (0.88 g, 3.85 mmol) in 40 mL anhydrous THF was treated with BOC 2 O (1.73 g, 7.72 mmol) and the reaction mixture was warmed at reflux (65 for 18 h.

The solvents were removed under reduced pressure, and flash chromatography (SiO2, 3 10 cm, 10% EtOAc/hexane) provided -12pure 104 as a yellow oil (0.96g, 'H NMR (CDC13, 250 MHz) 7.72 (br s, 1H), 6.86 (br s, 1H), 5.18 2H), 3.75 (s, 3H), 3.67 3H), 3.51 3H), 2.19 3H), 1.50 9H); NMR (CDC1 3 100 MHz) 5 152.7, 146.4, 143.6, 141.6, 127.8, 124.8, 105.3, 95.5, 80.4, 60.5, 60.4, 56.4, 28.3, 9.5; IR (film) v, 3437, 3341, 2977, 2935, 1731, 1519, 1454, 1422, 1397 cm-1; FABHRMS (NBA/CsI) m/z 460.0723 (C, 1 H,2NOs Cs* requires 460.0736).

[N-(tert-Butyloxycarbonyl)amino] (dimethoxy)-6-(iodo) ethoxymethoxy)-3-methyl benzene (10g). A solution of 104 (0.55 g, 1.67 mmol) in 6.6 mL anhydrous THF was cooled to (C and treated with TMEDA (0.94 mL, 6.18 mmol) followed by n-BuLi (2.5 mL of a 2.5 M solution in hexane, 6.18 mmol) in a slow dropwise manner. The resulting gold solution stirred for 2 h at -25 The reaction mixture was treated with l-chloro-2-iodoethane (0.45 mL, 6.18 mmol) and stirred for min at 25 The reaction was diluted with H20 (50 mL) and extracted with Et20 (3 30 mL), and the combined organic extracts were washed with saturated aqueous NaC1, dried (NaSO 4 and concentrated under reduced pressure. Flash chromatography (Si02, 2.5 10 cm, 20% EtOAc/hexane) yielded llg (560 mg, 74%) as a colorless oil: 'H NMR (CDC13, 400 MHz) 6 5.99 (br s, 1H), 5.10 2H), 3.75 3H), 3.69 3H), 3.65 3H), 2.17 3H), 1.49 9H); "C NMR (CDC13, 100 MHz) 5 153.6, 151.8, 150.5, 146.9, 129.3, 126.7, 99.1, 95.8, 80.6, 60.5, 60.3, 58.5, 28.3, 9.8; IR (film) 3321,-2975, 2936, 1722, 1485, 1455, 1390 cm-1; FABHRMS (NBA/CsI) m/z 585.9688

(C,

6

H

4 1INO 6 Cs requires 585.9703).

[N-(tert-Butyloxycarbonyl) 3 -chloro-2-propen-l-yl)amino]-2, 4-(dimethoxy)-6-(iodo)-5-(methoxymethoxy)- 3 -methylbenzene -13- (llg).

A solution of 10g (0.610 g, 1.34 mmol) in 13.4 mL anhydrous DMF was cooled to 0 and treated with NaH dispersion in oil, 121 mg, 4.03 mmol) in small portions. The resulting suspension was stirred for 15 min and treated with neat 1,3-dichloropropene (0.52 mL, 5.5 mmol) in a slow dropwise manner, followed by catalytic n-Bu 4 NI (50.0 mg, 0.13 mmol). The reaction mixture was warmed to 25 (C and stirred for 3 h. The reaction mixture was quenched with the addition of saturated aqueous NaHCO 3 (50mL), and the aqueous layer was extracted with EtOAc (3 30 mL). The combined organic extracts were washed with H 2 0 (4 50 mL), dried (Na 2

SO

4 and concentrated under reduced pressure. Flash chromatography (SiO 2 3 10 cm, 0-20% EtOAc/hexane gradient) yielded llg (0.681 g, 96%) as a colorless mixture of rotamers: IH NMR (CDC1 3 400 MHz) 2:1 rotamers 5 6.15-6.03 1H), 6.00-5.90 1H), 5.11-5.03 2H), 4.17-3.87 2H), 3.77 and 3.74 3H), 3.65 and 3.63 3H), 3.627 and 3.622 3H), 2.14 and 2.13 3H), 1.50 and 1.34 9H); 3 C NMR (CDC13, 100 MHz) rotamers 5 153.65 and 153.62, 152.8 and 152.1, 151.0 and 150.7, 147.0 and 146.7, 134.5 and 134.0, 129.5 and 129.0, 127.0 and 126.7, 121.0 and 120.6, 99.0, 97.7 and 97.3, 80.8 and 80.6, 60.5 and 60.4, 60.3 and 60.2, 58.5 and 58.4, 50.4, 48.8, 28.3 and 28.2, 9.9; IR (film) vmax 2973, 2936, 1704, 1456, 1366 cm-1; FABHRMS (NBA/CsI) m/z 659.9655 (C, 19

H

2 ClINOC Cs' requires 659.9626).

-14rN2 rYC SrN2 SOCI, 70 CH; Sr NH

H

2 N B O2s r Pd (PPh CO z OH 100 oC, 92% OR, EN n-BuOH. n-Bu 3

N

Z BnBr.NaH 210.R-H 170 0 C7421 OBn 100 OC.774% R Bn

RO

2 C NHBOC MeC 2 C

NHC

Nca. TsOH. 88%N OBn On LiOMe Z13. R n-Bu O 91% l-214, R Me 3 -Bromo-8-hydroxy-6-nitroquinoline (210).

A solution of 2-bromoacrolein (5 g, 37.0 mmol, 1 equiv) in 110 mL glacial acetic acid at 25 OC was titrated to the appearance of a faint reddish color with bromine (ca. 5.9 g, 37.0 mmol, 1 equiv). 2 -Hydroxy-4-nitroaniline (209, 5.7 g, 37.0 mmol, 1 equiv) was added, and the solution was gradually heated to 100 The solution was cooled to 25 (C after one hour. Filtering and neutralization of the precipitate with 1 M sodium phosphate buffer (pH 7, Na,HPO 4 -NaH 2

PO

4 afforded 9.2 g (9.95 g theoretical, 92%) of 210 as a light yellow solid: mp 240-241 1H NMR (CDC1 3 400 MHz) 5 8.93 J 2.0 Hz, 1H), 8.50 J 2.0 Hz, 1H), 8.23 J 2.2 Hz, 1H), 8.18 1H), 7.92 J 2.3 Hz, 1H); 3 C NMR (DMSO, 62.5 MHz) 155.1, 152.1, 146.3, 139.5, 139.1, 128.7, 119.1, 113.5, 105.1; IR (film) 3408 3089, 1587, 1553, 1519, 1476, 1389, 1350, 1297, 1263, 1210, 1133, 1079, 929, 931, 839, 804, 734, 633 ESIMS m/z 269 (M

C

9

H

3 BrO requires 269); Anal.

Calcd for C 9

H

3 BrO: C, 40.18; H, 1.87; N, 10.41. Found:

C,

40.21; H, 1.91; N, 9.98.

8 -(Benzyloxy)-3-bromo-6-nitroquinoline (211). A solution of 210 (13.7 g, 51 mmcl, 1 equiv) in anhydrous DMF (150 mL) was cooled to 4 (C under nitrogen and treated with KI (1.7 g, mmol, 0.2 equiv) and sodium hydride (60% dispersion in oil, 2.24 g, 56 mmol, 1.1 equiv). Benzyl bromide (7.3 mL, 6.1 mmol, 1.2 equiv) was added after 30 min and the reaction was allowed to warm to 25 After 24 h, the reaction volume was reduced by one-third in vacuo and EtOAc (200 mL) was added.

The reaction mixture was poured on H 2 0 (200 mL) and extracted with EtOAc (3 100 mL). The combined organic extracts were washed with saturated aqueous NaCl (1 40 ml), dried (Na 2

SO

4 and concentrated. Flash chromatography (SiO,, 5.5 20 cm, 100% CH 2 Cl 2 -hexane) afforded 211 (15.53 g, 18.32 g theoretical, as a yellow solid: mp 170 IH NMR (CDC1 3 400 MHz) 9.06 J 2.2 Hz, 1H), 8.44 J 2.2 Hz, 1H), 8.25 (d, J 2.2 Hz, 1H), 7:83 J 2.2 Hz, IH), 7.52 (app d, J 7.4 Hz, 2H), 7.38 2H), 7.32 1H), 5.47 2H); 13 C NMR (CDC1 3 62.5 MHz) 6 155.4, 153.3, 146.4, 140.3, 138.8, 135.0, 128.7 128.6, 128.4 127.5, 119.9, 115.1, 103.3, 71.4; IR (film) (m 3082, 3055, 2933, 2871, 1609, 1567, 1519, 1476, 1450, 1375, 1338, 1311, 1252, 1135, 1093, 976, 912, 842, 741 cm-1; FABHRMS (NBA/NaI) m/z 359.0040 (M

C

16 HBrN 2 03 requires 359.0031). Anal. Calcd for C, 6 HuBrN 2 0 3 C, 53.50; H, 3.09; N, 7.80. Found: C, 53.81; H, 3.23; N, 7.48.

8-(Benzyloxy)-3-bromo-6-N-(tert-butyloxycarbonyl)aminoquinolin e (212). A solution of 211 (200 mg, 0.56 mmol, 1 equiv) in EtOAc (1.1 mL) at 25 (C was treated with SnCl 2 -2H 2 0 (628 mg, 2.78 mmol, 5 equiv). The reaction mixture was heated to 70 (C under nitrogen until an orange slurry formed (ca. 0.5 h).

After cooling to 25 the reaction mixture was poured on ice and made basic with IN NaOH. The aqueous layer was filtered and extracted with EtOAc (3 15 mL). The combined organic layers were treated with saturated aqueous NaCl (l 10 mL), -16dried (Na 2

SO

4 and concentrated. The yellow solid was placed under vacuum for 0.5h and then dissolved in anhydrous dioxane mL) and treated with di-tert-butyl dicarbonate (490 mg, 2.25 mmol, 4.0 equiv) and triethylamine (156 AL, 1.12 mmol, equiv) The reaction mixture was warmed to 70 0 C under argon for one day. After cooling, the solvent was removed in vacuo. Chromatography (SiO 2 3 13 cm, 25% EtOAc-hexane) afforded 212 (179 mg, 240 mg theoretical, 74%) as a light yellow solid: mp 162 'H NMR (CDC13, 500 MHz) 5 8.77

J

2.0 Hz, 1H), 8.13 J 2.5 Hz, 1H), 7.47 (app d, J Hz, 2H), 7.42 J 2.0 Hz, 1H), 7.35 2H), 7.28 1H), 7.01 J 2.0 Hz, 1H), 6.61 1H), 5.37 2H), 1.51 (s, 9H); 13C NMR (CDC1 3 125 MHz) 5 154.9, 152.4, 148.3, 137.9, 136.3, 136.2, 135.4, 131.0, 128.6 128.0, 127.3 (2C), 118.6, 104.7, 103.6, 81.1, 70.9, 28.3 IR (film) (max 3354, 2971, 2919, 1807, 1766, 1724, 1621, 1450, 1367, 1310, 1253, 1217, 1160, 1123, 1061, 843, 771, 699, 657 cm-1; FABHRMS (NBA/CsI) m/z 429.0825 (M

C

21

H

21 BrN203 requires 429.0814).

n-Butyl8- (benzyloxy) (tert-butyloxycarbonyl) aminoquinoline 3 -carboxylate (213).

A solution of 212 (4.4 g, 10.1 mmol, 1 equiv) in 85 mL n-BuOH was cooled to -78 (C and degassed under vacuum.

Pd(PPh 3 4 (1.2g, 1.0 mmol, 0.1 equiv) and n-Bu 3 N (2.9 mL, 12.1 mmol, 1.2 equiv) were added and the solution was purged with nitrogen. The reaction mixture was then flushed with carbon monoxide and then slowly heated to 100 (C under a CO atmosphere. Upon complete reaction (ca. 12 50 mL H 2 0 and mL saturated aqueous

NH

4 C1 were added. The organic layer was separated and the aqueous layer was extracted with EtOAc (3 mL). The combined organic layers were washed with saturated aqueous NaCl (1 40 mL), dried (NaSOJ) and -17concentrated. Chromatography (SiO 2 5.5 20 cm, EtOAc-hexane) afforded 213 (3.55 g, 4.55 g theoretical, 78%) as a yellow solid: mp 135-136 'H NMR (CDC1 3 400 MHz) 9.31 J 2.0 Hz, 1H), 8.66 J 2.1 Hz, 1H), 7.61 (d, J 1.8 Hz, 1H) 7.49 (app d, J 7.4 Hz, 2H) 7.35 (app t, J 7.2 Hz, 2H) 7.29 1H) 7.11 J 2.1 Hz, 1H), 6.64 (br s, 1H), 5.39 2H), 4.38 J 6.6 Hz, 2H), 1.78 (m, 2H), 1.52 9H), 1.49 2H, buried under 1.52 ppm), 0.98 J 7.4 Hz, 3H); 13C NMR (acetone d 6 100 MHz) 5 165.7, 155.7, 153.5, 146.8, 139.8, 139.5, 137.8, 137.7, 129.5, 129.1 128.6, 128.4 (2C) 124.8, 107.1, 106.9, 80.4, 71.2, 65.5, 31.3, 28.3 19.8, 13.9. IR (film) 3222, 3049, 2958, 2930, 2876, 1717, 1617, 1544, 1503, 1430, 1362, 1271, 1239, 1157, 1065 cm- 1 FABHRMS (NBA/CsI) m/z 451.2249 (M H-,

C

26

H

30

N

2 0 5 requires 451.2233).

Methyl-8- (benzyloxy) 6 (tert-butyloxycarbonyl) aminoquinoline -3-carboxylate (214). A solution of 213 (2.9g, 6.4 mmol, equiv) in 70 mL MeOH was cooled to 4 (C under nitrogen and treated with LiOMe (275 mg, 7.1 mmol, 1.1 equiv). The reaction mixture was allowed to warm to 25 (C after 20 min.

Upon complete reaction (ca. 1.5 100 mL H 2 0 was added. The organic layer was separated and the aqueous layer was extracted with EtOAc (3 30 mL) The organic layers were combined, washed with saturated aqueous NaCl (1 30 mL), dried (Na 2 SO) and concentrated. Chromatography (SiO 2 5 19 cm, 25 30% EtOAc-hexane) afforded 214 (2.39 g, 2.63 g theoretical, 91%) as a yellow solid: mp 173-174 'H NMR (CDC13, 400 MHz) 5 9.29 J 2.0 Hz, 1H), 8.66 J 2.1 Hz, 1H), 7.56 J 1.8 Hz, 7.45 2H), 7.30 (m, 2H), 7.25 1H), 7.18 J 2.0 Hz, 1H), 6.85 1IH), 5.36 2H), 3.98 3H), 1.50 9H); 3 C NMR (CDC1 3 125 MHz) 5 165.9, 154.6, 152.6, 146.9, 138.6, 137.9, 137.8, 136.0, -18- 128.5, 128.4 127.9, 127.3 (2C) 123.8, 106.5, 105.5, 80.8, 70.8, 52.4, 28.2 (3C) IR (film) (max 3333, 3241, 2974, 1723, 1621, 1580, 1539, 1497, 1431, 1390, 1364, 1277, 1231, 1164, 1126, 1103, 1062, 1000, 882, 846, 795, 749, 697, 662 cm-1; FABHRMS (NBA/CsI) m/z 409.1773 (M

C

23

H

24

N

2 0 5 requires 409.1763).

Methyl 8- (benzyloxy) (tert-butyloxycarbonyl) amino] iodoquinoline-3-carboxylate A solution of 214 (2.13 g, 5.2 mmol, 1 equiv) :in 85 mL of a 1:1 mixture of THF-CH 3 0H was cooled to 4 (C and treated with TsOH (or H 2 S0) in 0.5 mL THF. N-Iodosuccinimide (1.4 g, 6.2 mmol, 1.2 equiv) in 10 mL THF was then slowly added over min. After 1.5 h, the reaction mixture was warmed to 25 (C and then stirred 45 h. Upon complete reaction, 100 mL saturated aqueous NaHCO,, 100 mL EtO, and 100 mL H 2 0 were added. The organic layer was separated and the aqueous layer was extracted with Et20 (3 50 mL) and EtOfAc (1 50 mL). The organic layers were combined, washed with saturated aqueous NaHCO 3 (1 50 mL) and saturated aqueous NaCl (1 50 mL), dried (Na 2

SO

4 and concentrated. Chromatography (SiO 2 5 19 cm, hexanes then 30% EtOAc-hexane) provided 10c (.2.34 g, 2.78 g theoretical, 84%, typically 80-88%) as a yellow solid: mp 182-183 'H NMR (CDC13, 400 MHz) 5 9.27 J 1.9 Hz, 1H), 8.96 J 1.9 Hz, 1H), 8.40 1H), 7.58 2H), 7.36 2H), 7.27 1H), 7.26 1H), 5.43 2H), 4.01 3H), 1.55 9H); "C NMR (CDC1 3 62.5 MHz) 5 165.4, 155.0, 152.3, 147.6, 141.9, 139.8, 139.7, 135.8, 129.6, 128.5 128.1, 128.0 125.1, 105.6, 81.7, 78.4, 71.1, 52.6, 28.2 IR (film) (mx3384, 2974, 1723, 1595, 1554, 1498, 1431, 1400, 1359, 1328, 1262, 1226, 1149, 995, 754 cm- 1 FABHRMS (NBA/CsI) m/z 535.0743 (M C 23

H

23

IN

2 0s requires -19- 535.0730).

H

H

NCOPh NIS /H N 2 M NaOMe/MeOH PN 0 0 0C.RT IN CM, 10min RT, PhOC 08n Pd 08n 89%

SOS

2M NaOMe/ I H NaD, ~COPh MeOH NN. N 0CM, 10mm N R7, then R 08n R7, 82% Bd On E/Z-1,3-dichloropropene, 16hr, RT, Di-t-Boc-Dicarbonate, DMAPf R H .31 10 66% 0CM, 3Omiri, RT, 93% SCC Z11

CI

N N'u1-Dibenzoyl-5.amino-.7-(benzyloxy)- 4 -iodoindole (305).

The above indole (118mg, Q.26mmcl) was stirred in THF (lm.L) and toluenesulfonic acid (26mg, 0.13mmol) was added. The solution was cooled to 0 0 C and N-iodosuccinimide (71mg, 0. 3l2mmol) in THF .(lmL) was added. The reaction was allowed to warm to 25 0 C over 1 hr. After l6hr, a further portion of N-iodosuccinimide (15mg, O.OG5mmol) was added and the reaction was stirred for a further 24hr. Saturated sodium bicarbonate solution (lmL) and water (4mL) were then added and the resulting mixture was extracted with chloroform (3 x 5mL).- The organic layers were combined, dried (MgSO,) and volatiles were removed under reduced pressure. The residue was purified by flash column chromatography (silica, ethyl acetate/hexane 3:7, x 15cm) and crystallized from ethyl acetate to give the expected product (305) as a yellow solid (67mg, 45%) 1 H NMR 6 (ppm) (CDC1 3 8. 15 Cs, IH, NH) 7. 99 2H Hz) 7. 64 (d, 2H, J Hz), 7.58-7.47 Cm, 4H, ArH), 7.30-7.17 Cm, SH, ArH), 6.58 Cd, 1H, J 3.6Hz), 4.92 2H). 13C NNR 5(ppm) 168, 165.5, 147.3, 136.2, 135.8, 134.9, 134.7, 134.1, 132.2, 132.1,129.8, 129.6, 129.5, 129.0, 128.4, 128.3, 128.2, 127.8, 127.7, 127.1, 127.0, 122.4, -110.6, 102.1, 71.9, 70.6 IR (neat) vmax. 3058, 1703, 1678, 1598, 1332, 1279, 1237, 695 crrv' Mass Spectrum (FAB, NAB/CsI) 705 Ns-Benzoyl-5-aznino-7- (benzyloxy) -4-iodoindole (310).

The above iodo-compound (305) (193mg, 0.34mmol) was stirred in dichioromethane C1OmL) Sodium methoxide in methanol CO.523mL, 1.O4MM0l) was added and the solution was stirred at RT for 10 min. Water (5OmL) and ethyl acetate (S0mL) were added and organic layer was separated, dried CMgSo,) and evaporated to give the crude product.

Chromatography (2 x 15cm SiO., ethyl acetate/hexanes 1:3) gave the pure compound (142mg, Rf 0.2 CSiO,,, ethyl acetate/hexanes 1: 3) as a colourless solid: 1H NMR CCDC1 3 400MHz) 8.61 Cs, 1H, NH), 8.31(s, 1H, NH), 8.05 1H), 8.00 Cd, 1H, J 6.8Hz) 7.53 Cm, 5H) 7.37 Cm, 3H) 7.21 Cdd, 1H, J 2.8, 1.4Hz), 6.43 Cdd, 1H, J 2.7, 1.2Hz) 5.28 Cs, 2H); IR (neat) va, 3290, 3010, 1658, 1573, 1535, 1355 cm'1; FARHRMS CNBA-CsT) m/z 600.9408.(M

C

2 ,H1 7

IN

2 0 2 requires 600.9389).

N

5 -Benzoyl-N, 1-di- (tert-butoxycarbonyl) -5-amino-7 Cbenzyloxy) 4-iodoindole (311).

Di-tert-butyl dicarbonate (687mg, 3.16mmol) and DMAP (128mg, l.O4mmol) were added to a stirred solution of compound 310 (255mg, 0.54mmol) in dichloromethane (6mL) After 30mmn at RT the solution was directly subjected to chromatography (2 x SiO., ethyl acetate/hexane 1:4) to give the pure product -21- (390mg, 93%) as a colorless oil (Rf 0.80, SiC 2 ethyl acetate/hexane 1: 3) 'H NNR (CDC.

3 400MHz) 7. 81 2H, J 6. 9Hz) 7. 55 Cd, 1H, J 3. 6Hz) 7. 45 (in, SH) 7. 31 (in, 3H) G. 82 1H) 6.57 1H, J 3.6Hz) 5.15 Cs, 2H) 1. 47 (s, 9H), 1.22 9H); FAB3HRMS CNBA-CsI) zn/z 801.0402 (M +Cs+,

C

32

H

3 3

IN

2 0 5 requires 801.0438)

N

5 1-Di- (tert-butoxycarbonyl) -5-antino-7- (benzyloxy) -4-iodoindo1 e Sodium methoxide in methanol (2M, 0.224mL, 0.44mrnol) Was added to a stirred solution of compound (311) (150mg, 0.22mmol) in dichioromethane (5mM After 10 min at RT, water and ethyl acetate (25mL) were added and the organic layer was separated. The aqueous layer was extracted with ethyl acetate C25mL) and the combined organic layers were dried CMgSO,) and concentrated. Chromatography (2 x SiO 2 ,gradient elution ethyl acetate/hexanes 1:9 to ethyl acetate/hexanes 1:3) gave the pure compound (102mg Rf 0.8 (SiC 2 ethyl acetate/hexanes 1:3) as a colourless oil: IH NMR (CDC1 3 400MHz) 7. 83 (br s, 1H, NH) 8. 00 Cd, 1H, J 6. 8Hz) 7. 53 Cm, 3H (1H 2H) 7.37 Cm, 3H) 6. 84 Cbr s, 1H), 6.45 Cd, 1N, J 3.4Hz) 5.21 2H) 1. 54 9H) 1. 45 (S, 9H) ;IR (neat) 3395, 2977, 1759, 1727, 1603, 1577, 1517, 1367, 1346, 1228, 1154, 1111 cm-1; FABHRMS (NBA-CsI) m/z 697. 0176 CM C 2

,H

2 9 IN.0 5 requires 697. 0176).

N

5 (3-Chloro-2-propen-1-yl)

-N

5 1-di- ((tere-butyloxy) carbonyl) -aniino-7- (benzyloxy) -4-iodoindole (lid).

Sodium hydride (22mg, 0.54mmo1, 3eq, 60% dispersion) was added to a stirred solution of compound Cl0d) (100mg, 0.l18mmol) in DMF (5mL) After 15 min at RT, E/Z-1,3-dichloropropene (0.025mL, 0.27mnol) was added. The -22solution was stirred at Rt for lhr. Water (50mL) and ethyl acetate (50mL) were then added and the organic layer was separated. The aqueous layer was extracted with ethyl acetate and the combined organic layers were dried (MgSO 4 and concentrated. Chromatography (2 x 15cm SiO,, ethyl acetate/hexanes 1:9) gave the pure compound (75.4mg 66%) as a mixture of E and 2 isomers: 'H NMR (CDC1 3 400MHz) major rotamer 7.47 (br s, 1H), 7.36 5H), 6.53 1H), 6.00 (m, 2H), 5.17 2H), 4.51 1H), 4.11 1H), 1.54 9H), 1.25 9H); IR (neat) Vm,, 2976, 1759, 1701, 1630, 1570, 1367, 1157 cm-1; FABHRMS (NBA-CsI) m/z 771.0125 (M+Cs C2,H 2 ClIN20 5 requires 771.0099) Throughout this specification and the claims.which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

The reference to any prior art in this specification is not, and should not be taken as, an acknowledgment or any form or suggestion that that prior art forms part of the common general knowledge in Australia.

Claims (8)

11-08-'06 10:43 FROM- T-983 P011/025 F-815 klO#Iv Da0spe0s02,2ilam I Ir aw -I I lma 0 -23- (c 3 THE CLAIMS DEFINING THE INVENTION ARE AS FOLLOWS: 1. A method for the synthesis of a dihydroindold C-ring of a CC-1065 duocarmycin analog wherein the method comprises 00 5 the steps of: C Step A: allylating an aryl halide with 1,3- en dichloropropene in the presence of a catalytic amount 1 of n-tetrabutylammonium iodide to form a vinyl C( 10 chloride; then Step B: cyclizing the vinyl chloride of said step A under conditions using tribuytyl tin hydride, catalytic AIBN and toluene as the solvent for forming the dihydroindole C-ring of the CC-1065 duocarmycin analog. 2. A method according to claim 1 wherein the aryl halide is selected from one of 10a-10g to afford the respective dihydroindole from 12a-g as depicted in Figure 6. 3. A compound represented by the following structure: "C0 2 Me N BOC. COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:43 FROM- T-983 P012/025 F-815 PVWEIPDBiI Iraa IlW22I Ips.dl IEMW -24- 4. A compound represented by the following structure: 5. A compound represented by the following structure: O Me N BOC. OMe 6. A compound represented by the following structure: CO 2 Me N IOC Bno NH8OC. 7. A compound represented by the following structure: COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:43 FROM- T-983 P013/25 F-815 iDi lsAo"eINDEv 8. A compound represented by the following structure: HN Br B NHBOC. 9. A compound represented by the following structure: OMOM MeO I Me# NHBOC OMe A compound represented by the following structure: BOC 11. A compound represented by the following structure: CO 2 Me an N BOC COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:44 FROM- T-983 P014/025 F-815 -26-

12. A compound represented by the following structure: BOC.

13. A compound represented by the following structure: HN Br l BOC

14. A compound represented by the following structure: OMOM CI M eooN I I I Me N OMe O 0 0 A compound represented by the following structure: BOC. COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11 11-08-'06 10:44 FROM- T-983 P015/025 F-815 :eeKlnaaiea iinpsu2 t aIgs-doiiM m -27-

16. A compound represented by the following structure: BOC

17. A compound represented by the following structure: OMOM Cl Mo Me N OM BOC OMe

18. A method according to claim 1 substantially as hereinbefore described, exemplified or illustrated.

19. A compound prepared by the method of claim 1. DATED this 11th day of August, 2006 The Scripps Research Institute By DAVIES COLLISON CAVE Patent Attorneys for the Applicant COMS ID No: SBMI-04430837 Received by IP Australia: Time 10:55 Date 2006-08-11