An Update on 2,5-Diketopiperazines from Marine Organisms

2,5-Diketopiperazines (2,5-DKPs) are an important category of structurally diverse cyclic dipeptides with prominent biological properties. These 2,5-DKPs have been obtained from a variety of natural resources, including marine organisms. Because of the increasing numbers and biological importance of these compounds, this review covers 90 marine originated 2,5-DKPs that were reported from 2009 to the first half-year of 2014. The review will focus on the structure characterizations, biological properties and proposed biosynthetic processes of these compounds.


Introduction
2,5-Diketopiperazines (2,5-DKPs) are important cyclodipeptides derived from the "head to tail" cyclization of two α-amino acids. These molecules with the double lactam core structure of 2,5-DKPs, have previously been isolated from a variety of natural resources, including marine organisms. These small, conformationally rigid, chiral templates have multiple sites in 2,5-DKPs for the structural elaboration of diverse functional groups with defined stereochemistry. These characteristics not only enable them to show a broad range of biological activities [1], but also allow the development of the drug-like physicochemical properties. The structures, reactions, medicinal chemical properties and potential therapeutic applications of 2,5-DKPs, particularly that with the interesting biological activities have previously been reviewed [2,3]. However, 2,5-DKPs belong to a relatively unexplored category of the bioactive cyclic peptides that may hold a great promise for the potential medicinal use in the future. Our previous review [1] focused on the marine-derived 2,5-DKPs, covering their structures, names, biological studies and proposed biosynthetic process. This review aims to summarize 90 marine organisms-derived 2,5-DKPs published from 2009 to the first half year of 2014. This update is taxonomically presented based on the origin of the isolation of these 2,5-DKPs.

Fungi from Mangrove Rhizosphere Soil Origin
The racemic spiroalkaloids, effusin A and dihydrocryptoechinulin D (shown here as one of the enantiomers, effusin A (50) and dihydrocryptoechinuline D (51), respectively) were obtained from the A. effuses (mangrove rhizosphere soil, Fujian, China). The racemates were subsequently resolved, and their absolute configurations were determined by the solution time dependent density function theory (TDDFT) and electronic CD (ECD) calculations. Effusin A (50) contains a spirobicyclic N,O-acetal moiety, which could be obtained by a domino ring-closure reaction between the substituted salicylaldehyde moiety in aspergin and the eneamide moiety of the DKP unit in neoechinulin B. In contrast, an enzyme-catalyzed regiospecific [4 + 2] Diels-Alder reaction produces the spirobicycle of dihydrocryptoechinuline D (51). The racemate of dihydrocryptoechinulin D inhibited the growth of P388 cells, and the (12R,28S,31S)-enantiomer 51 showed a selective, moderate inhibition of topoisomerase I [28]. Aspergilazine A (52), a DKP dimer consisting of two DKP units with a rare N-1 to C-6 linkage, was obtained from the A. taichungensis (mangrove root soil, Acrostichum aureum, source not given) [29]. The prenylated indole DKP alkaloid, dihydroneochinulin B (53) was isolated from the fungus A. effuses H1-1 (mangrove rhizosphere soil, Fujian, China) [30].

Fungi from Mollusk Origin
The fermentation of Aspergillus sp. (mollusk Mytilis edulis galloprovincialis, Noto Penninsula, Sea of Japan) yielded the notoamides O-R (70-73). Notoamide O (70) is noteworthy as the compound consists of a novel hemiacetal/hemiaminal ether moiety, which represents an unusual branch point for the oxidative modification of other members in the family of the prenylated indole alkaloids in the biogenetic pathway (Scheme 4) [42]. The structure of notoamide Q (72) has been revised [42]. The whole genome sequencing of Aspergillus MF 297-2 (mollusk M. edulis galloprovincialis, Noto Penninsula, Japan Sea) [52] led to the identification and characterization of the biosynthetic gene cluster for stephacidins [53] and the notoamide alkaloids [43,52]. Scheme 4. Postulated biosynthetic pathway for 70 [42].

Fungi from Other Origins
Notamide E (74) was isolated from the culture of Aspergillus sp. (mussel Mytilus edulis, Noto Peninsula, Sea of Japan) [43]. The compound (74) has been synthesized prior to its isolation from the natural source, and it was proposed [44] to be an advanced precursor to notoamides A-D. The biosynthetic studies of the producing organism indicated that notoamide E (74) was a short-lived metabolite. The feeding experiments utilizing synthetic, 13 C-labelled (74) demonstrated the incorporation of notoamide E into notoamides C [43], D [45] and 3-epi-notamide C [44]. These studies also produced three minor alkaloids, notamides E2-E4 (75-77) [45]. Further investigation of the same culture of Aspergillus that yielded notamides A-D [45] led to the isolations of notamides L-N (78-80) [46]. In a direct and targeted gene manipulation experiment, the provision of synthetic N-alkyl tryptophan to a prenyltransferase-deficient mutant of a cyclomarin/cyclomarazine-producing S. arenicola led to the discovery of some novel derivatives, cyclomarazines M (81) and P (82) [47]. Two central pathway enzymes, which catalyzed both the normal and reverse prenyltransfer reactions, were characterized. The study also established the early steps of the biosynthetic procedure of prenylated indole alkaloid structure, including the production of notoamide S (83) [48]. Notoamide S (83) has been synthesized via N-Fmoc proline coupling with a 6-hydroxy-7-prenyl-2-reverse prenyl tryptophan derivative [49]. The spirocyclic DKP alkaloid, spirotryprostatin F (84) was isolated from the A. fumigatus (soft coral Sinularia sp., Kunashir Is., Kuril Islands), and it showed a stimulatory phytoregulatory activity in the study [50]. An indolyl DKP compound, penilloid A (85) was isolated from two marine derived fungi Penicillium sp. and A. sydowii [51].

Gorgonian
The isolation of the gorgonian Menella kanisa collected from Beibu Gulf led to the identification of menazepine A (89) (Figure 12) [57].

Conclusions
2,5-DKPs are ubiquitous in nature. They have previously been isolated from bacteria, fungi, marine invertebrates and higher organisms [1,2]. Although these DPK derivatives have been known since the early 20th century, they only recently draw significant interests because of the diverse range of their biological activities [1], including the disruption of the biofilm formation through modulation of bacterial quorum sensing and their role in an interkingdom cell-cell signaling [42]. The increasing numbers of naturally occurring bioactive 2,5-DKPs have been obtained from various marine organisms, and the studies on these 2,5-DKPs have been the focus of many recent studies because of their potent biological activities. To date, more than 200 2,5-DKPs have been isolated from a diverse range of marine organisms, particularly marine microorganisms. Some of these 2,5-DKPs exhibited various bioactivities, such as cytotoxicity on cancer cell lines, anti-microbial and anti-inflammatory properties [1]. From 2009 to the first half-year of 2014, the main natural source of DKPs isolated from marine organisms is marine microorganisms, accounting for 94% ( Figure 14). Many studies have performed on these bacteria, fungi and actinomycete that produced 2,5-DKPs, and these marine microorganisms were isolated from sediments, algae, mangrove, sponges and mud ( Figure 15). It was indicated that 51% of the studied microorganisms were isolated from sediments as shown in Figure 15. Interestingly, the marine-derived fungi accounted for the largest part (84%) of total 2,5-DKPs that were isolated from marine microorganisms ( Figure 16). The interest in natural products from marine microorganisms, especially marine-derived fungi, has increased significantly in the last decade [59,60], which has led to the discoveries of more 2,5-DKPs from the marine-derived fungi than that from other marine organisms. Consequently marine microorganisms, especially fungi can be a promising source of these bioactive 2,5-DKPs. The discovery of these compounds from marine-derived fungi demonstrates that some gene clusters in fungi may have the ability to produce structurally diverse DKPs by the biosynthetic pathways, which may need further investigations.   as academic visitor at University of Bath (with Ian S Blagbrough), and thanks Heather Wyman-Pain, University of Bath, for proofreading.

Author Contributions
Ri-Ming Huang contributed in writing the manuscript. Cheng-Hai Gao conceived and designed the format of the manuscript. All the authors contributed in critical reading and discussion on the manuscript.

Conflicts of Interest
The authors declare no conflict of interest.