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Enediyne Natural Products: Biosynthesis and Prospect Towards Engineering Novel Antitumor Agents

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Enediyne Natural Products: Biosynthesis and Prospect Towards Engineering Novel Antitumor Agents Current Medicinal Chemistry, 2003, 10, 2317-2325 2317 Enediyne Natural Products: Biosynthesis and Prospect Towards Engineering Novel Antitumor Agents Ben Shen1,2*, Wen Liu1 and Koichi Nonaka1 1Division of Pharmaceutical Sciences and 2Department of Chemistry, University of Wisconsin, Madison, WI 53705, USA Abstract: This review gives a brief account on the current status of enediyne biosynthesis and the prospective of applying combinatorial biosynthesis methods to the enediyne system for novel analog production. Methods for cloning enediyne biosynthetic gene clusters are first reviewed. A unified paradigm for enediyne biosynthesis, characterized with (a) the enediyne PKS, (b) the enediyne PKS accessory enzymes, and (c) tailoring enzymes, is then presented. Strategies and tools for novel enediyne analog production by combinatorial biosynthesis are finally discussed. The results set the stage to decipher the molecular mechanism for enediyne biosynthesis and lay the foundation to engineer novel enediynes by combinatorial biosynthesis for future endeavors. Keywords: Biosynthesis, C-1027, calicheamicin, combinatorial biosynthesis, enediyne, polyketide synthase. INTRODUCTION categories according to the enediyne core structures. Members of the 9-membered enediyne core sub-category are Neocarzinostatin (NCS), the first member of the enediyne chromoproteins consisting of an apo-protein and the family of antitumor antibiotics, was originally discovered as enediyne chromophore, with N1999A2 (5 ) from a macromolecular antitumor antibiotic from the culture Streptomyces sp. AJ9493 as the only exception that was filtrates of a Streptomyces carzinostaticus strain in 1965 [1]. isolated as a chromophore alone [15]. The apo-protein acts as Although it became clear shortly after its discovery that all a stabilizer and specific carrier for the otherwise unstable biological activities of NCS resided in a nonprotein chromophore and its transport and interaction with target chromophore, the NCS chromophore structure (1) was not DNA. Due to their intrinsic reactivity, only three other elucidated until 1985 [2], revealing an unprecedented chromoprotein chromophore structures, in additional to 1 bicyclo[7,3,0]dodecadiynene system, i.e., the 9-membered and 5, are currently known—kedarcidin (6) from enediyne core, decorated with an aminosugar and a naphthoic Actinomycete L585-6 [16], C-1027 (7) from Streptomyces acid moiety. However, this seminal work was globisporus [17-20], maduropeptin (8) from Actinomadura underappreciated in the subsequent two years, and it was the madurae [21]. Members of the 10-membered enediyne core discovery of the calicheamicins (2) from a Micromonospora sub-category are in general more stable, all of which were echinospora strain [3,4] and the esperamicins (3) from an isolated as discrete small molecules. In addition to 2, 3, 4, Actinomadura verrucosospora strain [5,6] in 1987 that other members of this sub-category whose structures have grabbed the attention of chemists and biologists alike to the been elucidated include dynemicin (9 ) from enediynes. In contrast to 1, structural elucidation of 2 and 3 Micromonospora chersina sp. nov. No. M965-1 [22,23] and unveiled a novel biocyclo[7,3,1]tridecadiynene system, i.e., namenamicin (10) from a marine ascidian Polysyncraton the 10-membered enediyne core, decorated with several lithostrotum species [24] (Figure 1). deoxysugar moieties. Since then, the enediyne family of Members of both sub-categories of enediynes share a natural products has been the focus of intense research common mechanism of action, despite their structural activity in the fields of chemistry, biology, and medical difference. The enediyne core undergoes an electronic sciences because of their highly unusual molecular rearrangement (Bergman or Myers rearrangement) to form a architectures, biological activities, and modes of actions [7- transient benzenoid diradicals that damage DNA by 13]. abstracting hydrogen atoms from the deoxyribose moiety on Over twenty enediyne natural products are currently both strands. Subsequent reactions of the resultant known, and new members of the enediyne family are deoxyribose carbon-centered radicals with molecular oxygen continuously to be discovered with the newest addition, the initiate a process that leads in both single-strand and double- shishijimicins (4) from a marine ascidian Didemnum strand DNA cleavage [7-13,25-27]. This novel mechanism of proliferum species, reported in early 2003 [14]. The DNA damage has important implications for the enediyne natural products could be classified into two sub- development of the enediynes into clinical anticancer drugs [25-36]. Although the natural enediynes have seen limited use as clinical drugs mainly because of substantial toxicity, *Address correspondence to this author at the Division of Pharmaceutical various polymer-based delivery systems or enediyne- Sciences, School of Pharmacy, University of Wisconsin-Madison, 777 Highland Ave., Madison, WI 53705, USA; Tel: +1-608-263-2673; Fax: antibody conjugates have shown great clinical success and/or +1-608-262-5345; E-mail: [email protected] promise in anticancer chemotherapy [8,10,12,13,37,38]. For 0929-8673/03 $41.00+.00 © 2003 Bentham Science Publishers Ltd. 2318 Current Medicinal Chemistry, 2003, Vol. 10, No. 21 Shen et al. example, the poly(styrene-co-maleic acid)-conjugated NCS harnessed and delivered onto the target tumor cells. A great was approved in Japan in 1993 and has been marketed since challenge will be to develop ways to make new enediynes 1994 for use against hepatoma [10]. A CD33 monoclonal for mechanistic and clinical studies. antibody (mAB)-calicheamicin conjugate was approved in Access to complex natural products such as the enediynes US in 2000 and has been marked under the trade name of and their analogs by total synthesis poses a monumental Mylotarg to treat acute myeloid leukemia [37]. Several challenge to synthetic chemists, and yet tour de force effort antiheptoma mAB-C-1027 conjugates have also been by synthetic chemists has led to the total syntheses of prepared and showed to display high tumor specificity and almost every member of the enediyne family of natural to exert a strong inhibitory effect on the growth of products as well as a myriad of analogs. Significant progress established tumor xenografts [38]. These examples clearly in developing the enediynes into clinical agents has been demonstrate that the enediynes can be developed into made towards (a) improving cancer cell specificity, (b) powerful drugs when their extremely potent cytotoxicity is developing efficient delivery systems to the tumor targets, O A H2C O OCH3 O O O N OC H3 O O O O H3C O H C 3 O O O (C H3 )2 N OH O R CH3 OH OH O CH3HN 22 O HO O Cl CH 3 N HO R = OH (7) NH2 (1) R = H (11) O H Cl O OH HO O O OH H3 CO OH CH O NH 3 HO CH3 (8) H3C i-PrO OH OH HO O H3CO N(CH ) OC H3 O OCH 3 2 3 NH H3C Cl O O Cl N OH O HOH2C O O O O OH (5) O O OH H3C O HO (6) H3C Biosynthesis and Prospect Towards Engineering Novel Antitumor Agents Current Medicinal Chemistry, 2003, Vol. 10, No. 21 2319 (Fig. 1). contd..... B SSSC H3 O NHCO2CH3 OH O CH HO O 3 O O CH3 O N S O OH H OCH3 H3COCH3 O NHEt IOCH3 H3C O (2) HO CH O 3 OH SSSC H3 O NHCO 2CH3 OH O CH HO O 3 O O CH3 N SCH O O OH H 3 OCH3 H3C O O NHPr-i O HO H3CO O (3) H3CO NH O OC H3 SSSCH3 CH3 O H NHCO2CH3 CO2H OH OH O HN O CH O HO O 3 O O CH3 O HO SC H3 OC H3 SCH3 OH OCH3 O NHPr-i OH OOH (10) (9) OH SSSCH3 O N NHCO 2CH3 N H CH3 HO O O O HO O SCH3 OCH3 O NHPr-i (4) Fig. (1). The chromophore structures of naturally occurring 9-membered enediyne chromoproteins: neocarzinostatin (1), C-1027 (7), maduropetin (8), kedarcidin (6), and N1999A2 (5) and as well as the engineered deshydroxy-C-1027 (11). (B) Structural I representation of naturally occurring 10-membered enediynes: calicheamicin r1 (2), esperamicin A1 (3), dynemicin A (9), namenamicin (10), and shishijimicin A (4). and (c) designing triggers and trapping the enediynes as modification of the natural products can only access to prodrugs [11-13]. However chemical total synthesis has very limited functional groups, often requiring multiple limited practical value for complex natural products such as protection and deprotection steps. the enediynes, and analog generation by chemical 2320 Current Medicinal Chemistry, 2003, Vol. 10, No. 21 Shen et al. Complementary to organic synthesis, genetic of genome of the antibiotic-producing actinomycetes), a manipulations of genes governing secondary metabolism— daunting task that is further complicated by the fact that an emerging technology also known as combinatorial most of these organisms contain multiple secondary biosynthesis—offer a promising alternative to preparing metabolite biosynthetic gene clusters [49-51]. complex natural products and their analogs biosynthetically [39-46]. Specific structural alteration in the presence of other functional groups can often be achieved, and the target dNDP-Glucose 4,6-Dehydratase as A Probe molecules will be produced by a recombinant organism that This strategy was exemplified by the cloning of the is amenable for large-scale fermentation, thereby lowering biosynthetic gene cluster for 7 from S. globisporus [52,53] the production cost and reducing the environment concern and should be applicable to gene clusters of both the 9- and associated with conventional chemical synthesis. The 10-membered enediynes. Except for 5 and 9, enediynes with success of this approach depends on (a) the cloning and known structures all contain at least one deoxyhexose genetic and biochemical characterization of the biosynthetic moiety. The biosynthesis of various deoxyhexoses shares a pathways of the target metabolites and (b) the development common key intermediate, 4-keto-6-deoxyglucose nucleoside of strategies, methods, and expedient tools for combinatorial diphosphate, the formation of which from glucose manipulation of natural product biosynthetic gene clusters.
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