Design and Synthesis of Two Cytotoxic Analogs of the Novel Pyrrolo[1 0 ,2 0 :1,2][1,4]diazepin [7,6-b]indol-5(6H)-one Nucleus Andrew Tsotinis,  Margarita Vlachou, Konstantinos Kiakos, y John A. Hartley, yy and David E. Thurston y University of Athens, School of Pharmacy, Department of Pharmaceutical Chemistry, Panepistimiopolis-Zografou, GR-157 71, Athens, Greece y CR-UK Gene Targeted Drug Design Research Group, The School of Pharmacy, University of London, 29/39 Brunswick Square, London WC1N 1AX, U.K. yy CR-UK Drug-DNA Interactions Research Group, Department of Oncology, Royal Free & University College Medical School, UCL, 91 Riding House Street, London WIP 8BT, U.K. (Received February 28, 2003; CL-030170) The design and synthesis of the two cytotoxic derivatives 15 and 16 of the novel pyrrolo[1 0 ,2 0 :1,2][1,4]diazepin[7,6-b]in- dol-5(6H)-one nucleus is described. Readily available methyl 2- indolecarboxylates 5 and 6 are nitrosated with NaNO 2 in AcOH to give the analogs 7 and 8, which are then oxidized with KMnO 4 in aq. NaOH to provide the 3-NO 2 acids 9 and 10. These, in turn, are subjected to amidation with (2S)-pyrroli- dine-2-carboxaldehyde diethyl thioacetal in the presence of EDCI and HOBt and then to a 7-exo-trig cyclization reaction to give the target molecules 15 and 16. The new analogs were evaluated in the human leukemic K 562 cell line and were shown to have micromolar potency. The natural product anthramycin 1 and its naturally occur- ring (e.g. DC-81, 2) and synthetic pyrrolo[2,1-c][1,4]benzodi- azepine (PBD) congeners (e.g. 3 and 4) 1;2 have in recent years generated significant interest as the basis for novel types of anti- tumour agents (Figure 1). 3 The PBDs interact within the minor groove of DNA by forming a covalent bond between their elec- trophilic C11-position and the exocyclic C2-NH 2 moiety of a guanine residue. 4 Most of the known PBDs, including anthra- mycin, have a tricyclic skeleton which consists of an aromatic A-ring, a 1,4-diazepin-5-one B-ring bearing a N10-C11 imine functionality (or the equivalent) and a pyrrolidine C-ring. Through extensive Structure-Activity Relationship (SAR) stu- dies, each of these rings is known to play a distinct role in the interaction with DNA. For example, the (S) configuration of the C11a position at the B-C junction provides the molecules with the appropriate 3-dimensional shape to fit into the DNA minor groove. Similarly, the aromatic A-ring substituents can enhance binding of the PBD through the formation of hydrogen bonds to DNA bases. 4;5 It is known that the presence of a hydroxyl group at the C-9 position of the A-ring can lead to cardiotoxicity. 6 In an attempt to circumvent this, various analogs have been synthesized which bear nitrogen-containing aromatic heterocycles (e.g. 3) in place of the usual aromatic A-ring. We report here the design and synthesis of the first indolic PBD analogs 15 and 16 (Scheme 1). The indole nucleus was chosen because it is known to increase minor groove binding affinity without significantly affecting sequence selectivity. 7 Furthermore, the methoxyl groups in the indole moiety were considered to be potential sources of hydrogen bonds. In the structures of the new tetra- cyclic analogs 15 and 16 the B and C-rings of the parent PBDs have been retained. The strategy for the synthesis of the target molecules 15 and 16 is shown in Scheme 1. Methyl 5-methoxy-2-indolecarboxy- late (5) was prepared from commercially available 5-meth- oxyindole-2-carboxylic acid by reaction with thionyl chloride N N O CONH 2 OCH 3 OH H 3 C H H N N O HO H CH 3 O 1: Anthramycin 2: DC-81 1 2 3 4 5 6 7 8 9 10 11 11a 10 11 11a N N N O H OCH 3 3 10 11 11a N N N N O H CH 3 O OCH 3 4 10 11 11a H A B C Figure 1. N H CH 3 O CO 2 CH 3 R 5: R = H 6: R = OCH 3 N H CH 3 O CO 2 CH 3 NO R 7: R = H (62%) 8: R = OCH 3 (67%) N H CH 3 O CO 2 H NO 2 R 9: R = H (68%) 10: R = OCH 3 (75%) a b c N H CH 3 O NH 2 N O CH(SCH 2 CH 3 ) 2 H R 13: R = H (79%) 14: R = OCH 3 (82%) N H CH 3 O N N O H R 15: R = H (60%) 16: R = OCH 3 (65%) e N H CH 3 O NO 2 N O CH(SCH 2 CH 3 ) 2 H R 11: R = H (61%) 12: R = OCH 3 (68%) d Scheme 1. Reagents and conditions: (a) NaNO 2 , AcOH, 2 h; (b) KMnO 4 , aq. NaOH (2M), 30 min; (c) (2S)-pyrrolidine-2- carboxaldehyde diethyl thioacetal, EDCI, HOBt, Et 3 N, CH 2 Cl 2 -DMF (2:1), 18h; (d) 10% Pd-C, H 2 (50atm), MeOH, 3.5h; (e) HgCl 2 , CaCO 3 , CH 3 CN-H 2 O (4:1), 8h. 512 Chemistry Letters Vol.32, No.6 (2003) Copyright Ó 2003 The Chemical Society of Japan