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For exploring structurally diverse metabolites and uniquely metabolic mechanisms, we systematically investigated the chemical constituents and putative biosynthesis of Janibacter sp. SCSIO 52865 derived from the deep-sea sediment based on the OSMAC strategy, molecular networking tool, in combination with bioinformatic analysis. As a result, one new diketopiperazine ( 1 ), along with seven known cyclodipeptides ( 2 - 8 ), trans -cinnamic acid ( 9 ), N -phenethylacetamide ( 10 ) and five fatty acids ( 11 - 15 ), was isolated from the ethyl acetate extract of SCSIO 52865. Their structures were elucidated by a combination of comprehensive spectroscopic analyses, Marfey's method and GC-MS analysis. Furthermore, the analysis of molecular networking revealed the presence of cyclodipeptides, and compound 1 was produced only under mBHI fermentation condition. Moreover, bioinformatic analysis suggested that compound 1 was closely related to four genes, namely jat A-D, encoding core non-ribosomal peptide synthetase and acetyltransferase., Competing Interests: The authors declare no conflicts of interest.

New therapeutic options to combat the growing incidence of antimicrobial resistance are urgently needed. A 2015 publication reported the isolation and biological evaluation of two diketopiperazine natural products, cyclo(l-Trp-l-Arg) (CDP 2) and cyclo(d-Trp-d-Arg) (CDP 3), from an Achromobacter sp. bacterium, finding that the latter metabolite in particular exhibited strong antibacterial activity towards a range of wound-related microorganisms and could synergize the action of ampicillin. Intrigued by these biological activities and noting inconsistencies in the structural characterization of the natural products, we synthesized the four diastereomers of cyclo(Trp-Arg) and evaluated them for antimicrobial and antibiotic enhancement properties. The detailed comparison of spectroscopic data raises uncertainty regarding the structure of CDP 2 and disproves the structure of CDP 3. In our hands, none of the four stereoisomers of cyclo(Trp-Arg) exhibited detectable intrinsic antimicrobial properties towards a range of Gram-positive and Gram-negative bacteria or fungi nor could they potentiate the action of antibiotics. These discrepancies in biological properties, compared with the activities reported in the literature, reveal that these specific cyclic dipeptides do not represent viable templates for the development of new treatments for microbial infections.

An efficient method for the synthesis of functionalized peptidomimetics via multicomponent Ugi reaction has been developed. The application of trifluoroethanol (TFE) as a reaction medium provided desired products with good yields. Further, using the developed cyclisation reaction, the obtained peptidomimetics were transformed into the cyclic analogues (diketopiperazines, DKPs). The goal of the performed studies was to revised and compare whether the structure of the obtained structurally flexible acyclic peptidomimetics and their rigid cycling analogue DKPs affect antimicrobial activity. We studied the potential of synthesized peptidomimetics, both cyclic and acyclic, as antimicrobial drugs on model E. coli bacteria strains (k12, R2-R4). The biological assays reveal that DKPs hold more potential as antimicrobial drugs compared to open chain Ugi peptidomimetics. We believe that it can be due to the rigid cyclic structure of DKPs which promotes the membrane penetration in the cell of studied pathogens. The obtained data clearly indicate the high antibiotic potential of synthesized diketopiperazine derivatives over tested antibiotics.

The long-acting parenteral formulation of the HIV integrase inhibitor cabotegravir (GSK744) is currently being developed to prevent HIV infections, benefiting from infrequent dosing and high efficacy. The crystal structure can affect the bioavailability and efficacy of cabotegravir. However, the stability determination of crystal structures of GSK744 have remained a challenge. Here, we introduced an ab initio protocol to determine the stability of the crystal structures of pharmaceutical molecules, which were obtained from crystal structure prediction process starting from the molecular diagram. Using GSK744 as a case study, the ab initio predicted that Gibbs free energy provides reliable further refinement of the predicted crystal structures and presents its capability for becoming a crystal stability determination approach in the future. The proposed work can assist in the comprehensive screening of pharmaceutical design and can provide structural predictions and stability evaluation for pharmaceutical crystals.

2,5-diketopiperazines (DKPs) are cyclic dipeptides ubiquitously found in nature. In particular, cyclo(Phe-Pro), cyclo(Leu-Pro), and cyclo(Val-Pro) are frequently detected in many microbial cultures. Each of these DKPs has four possible stereoisomers due to the presence of two chirality centers. However, absolute configurations of natural DKPs are often ambiguous due to the lack of a simple, sensitive, and reproducible method for stereochemical assignment. This is an important problem because stereochemistry is a key determinant of biological activity. Here, we report a synthetic DKP library containing all stereoisomers of cyclo(Phe-Pro), cyclo(Leu-Pro), and cyclo(Val-Pro). The library was subjected to spectroscopic characterization using mass spectrometry, NMR, and electronic circular dichroism (ECD). It turned out that ECD can clearly differentiate DKP stereoisomers. Thus, our ECD dataset can serve as a reference for unambiguous stereochemical assignment of cyclo(Phe-Pro), cyclo(Leu-Pro), and cyclo(Val-Pro) samples from natural sources. The DKP library was also subjected to a biological screening using assays for E. coli growth and biofilm formation, which revealed distinct biological effects of cyclo(D-Phe-L-Pro).

Ten polyketide derivatives ( 1 - 10 ), including a new natural product named ( E )-2,4-dihydroxy-3-methyl-6-(2-oxopent-3-en-1-yl) benzaldehyde ( 1 ), and five known diketopiperazines ( 11 - 15 ), were isolated from the mangrove-sediment-derived fungus Aspergillus sp. SCSIO41407. The structures of 1 - 15 were determined via NMR and MS spectroscopic analysis. In a variety of bioactivity screening, 3 showed weak cytotoxicity against the A549 cell line, and 2 exhibited weak antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA). Compounds 3 , 5 , and 6 showed inhibition against acetylcholinesterase (AChE) with IC 50 values of 23.9, 39.9, and 18.6 μM. Compounds 11 , 12 , and 14 exhibited obvious inhibitory activities of lipopolysaccharide (LPS)-induced nuclear factor- κ B (NF- κ B) with IC 50 values of 19.2, 20.9, and 8.7 μM, and they also suppressed RANKL-induced osteoclast differentiation in bone marrow macrophages cells (BMMCs), with the concentration of 5 μM. In silico molecular docking with AChE and NF- κ B p65 protein were also performed to understand the inhibitory activities, and 1 , 11 - 14 showed obvious protein/ligand-binding effects to the NF-κB p65 protein.

Cyclodipeptides (CDPs) or 2,5-diketopiperazines (DKPs) can exert a variety of biological activities and display pronounced resistance against enzymatic hydrolysis as well as a propensity towards self-assembly into gels, relative to the linear-dipeptide counterparts. They have attracted great interest in a variety of fields spanning from functional materials to drug discovery. This concise review will analyze the latest advancements in their synthesis, self-assembly into gels, and their more innovative applications.

Previous studies revealed the potential of Labrenzia aggregata USBA 371 to produce cytotoxic metabolites. This study explores its metabolic diversity and compounds involved in its cytotoxic activity. Extracts from the extracellular fraction of strain USBA 371 showed high levels of cytotoxic activity associated with the production of diketopiperazines (DKPs). We purified two compounds and a mixture of two other compounds from this fraction. Their structures were characterized by 1D and 2D nuclear magnetic resonance (NMR). The purified compounds were evaluated for additional cytotoxic activities. Compound 1 (cyclo (l-Pro-l-Tyr)) showed cytotoxicity to the following cancer cell lines: breast cancer 4T1 (IC 50 57.09 ± 2.11 µM), 4T1H17 (IC 50 40.38 ± 1.94), MCF-7 (IC 50 87.74 ± 2.32 µM), murine melanoma B16 (IC 50 80.87 ± 3.67), human uterus sarcoma MES-SA/Dx5 P-pg (-) (IC 50 291.32 ± 5.64) and MES-SA/Dx5 P-pg (+) (IC 50 225.28 ± 1.23), and murine colon MCA 38 (IC 50 29.85 ± 1.55). In order to elucidate the biosynthetic route of the production of DKPs and other secondary metabolites, we sequenced the genome of L. aggregata USBA 371. We found no evidence for biosynthetic pathways associated with cyclodipeptide synthases (CDPSs) or non-ribosomal peptides (NRPS), but based on proteogenomic analysis we suggest that they are produced by proteolytic enzymes. This is the first report in which the cytotoxic effect of cyclo (l-Pro-l-Tyr) produced by an organism of the genus Labrenzia has been evaluated against several cancer cell lines.

The first total syntheses of izenamides A, B, and C, which are depsipeptides inhibitor of cathepsin D, were accomplished. In addition, the stereochemistry of izenamide B was confirmed by our syntheses. The key features of our synthetic route involve the avoidance of critical 2,5-diketopiperazine (DKP) formation and the minimization of epimerization during the coupling of amino acids for the target peptides.

The glyoxylate cycle is a sequence of anaplerotic reactions catalyzed by the key enzymes isocitrate lyase (ICL) and malate synthase, and plays an important role in the pathogenesis of microorganisms during infection. An icl -deletion mutant of Candida albicans exhibited reduced virulence in mice compared with the wild type. Five diketopiperazines, which are small and stable cyclic peptides, isolated from the marine-derived Streptomyces puniceus Act1085, were evaluated for their inhibitory effects on C. albicans ICL. The structures of these compounds were elucidated based on spectroscopic data and comparisons with previously reported data. Cyclo(L-Phe-L-Val) was identified as a potent ICL inhibitor, with a half maximal inhibitory concentration of 27 μg/mL. Based on the growth phenotype of the icl -deletion mutants and icl expression analyses, we demonstrated that cyclo(L-Phe-L-Val) inhibits the gene transcription of ICL in C. albicans under C 2 -carbon-utilizing conditions.

Three new thiodiketopiperazines ( 1 ⁻ 3 ), along with two known analogues ( 4 and 5 ), were isolated from the fermentation broth of Penicillium crustosum . Their structures were elucidated through extensive spectroscopic analysis and the absolute configurations of new compounds were determined by Mosher ester analysis and calculated ECD spectra. Compound 4 and 5 have the activity to promote the gastrointestinal motility of zebrafish via acting on the cholinergic nervous system.

The organic extract of liquid cultures of the marine-derived Penicillium sp. was investigated. Fractionation of the extracts of the fungus led to the purification and identification of two new compounds, penicillatides A ( 1 ) and B ( 2 ), together with the previously reported cyclo( R -Pro -S -Phe) ( 3 ) and cyclo( R -Pro -R -Phe) ( 4 ). The structures of compounds 1 - 4 were assigned by extensive interpretation of their NMR and high-resolution mass spectrometry (HRMS). The antiproliferative and cytotoxic activities of the compounds against three human cancer cell lines as well as their antimicrobial activity against several pathogens were evaluated. Compounds 2 - 4 displayed variable cytotoxic and antimicrobial activities., Competing Interests: The authors declare no conflict of interest.

Cyclodipeptides, called 2,5-diketopiperazines (2,5-DKPs), are obtained by the condensation of two amino acids. Fungi have been considered to be a rich source of novel and bioactive cyclodipeptides. This review highlights the occurrence, structures and biological activities of the fungal cyclodipeptides with the literature covered up to July 2017. A total of 635 fungal cyclodipeptides belonging to the groups of tryptophan-proline, tryptophan-tryptophan, tryptophan-Xaa, proline-Xaa, non-tryptophan-non-proline, and thio-analogs have been discussed and reviewed. They were mainly isolated from the genera of Aspergillus and Penicillium. More and more cyclodipeptides have been isolated from marine-derived and plant endophytic fungi. Some of them were screened to have cytotoxic, phytotoxic, antimicrobial, insecticidal, vasodilator, radical scavenging, antioxidant, brine shrimp lethal, antiviral, nematicidal, antituberculosis, and enzyme-inhibitory activities to show their potential applications in agriculture, medicinal, and food industry., Competing Interests: The authors declare no conflict of interest.

Diketopiperazines can be generated by non-enzymatic cyclization of linear dipeptides at extreme temperature or pH, and the complex medium used to culture bacteria and fungi including phytone peptone and trypticase peptone, can also produce cyclic peptides by heat sterilization. As a result, it is not always clear if many diketopiperazines reported in the literature are artifacts formed by the different complex media used in microorganism growth. An ideal method for analysis of these compounds should identify whether they are either synthesized de novo from the products of primary metabolism and deliver true diketopiperazines. A simple defined medium ( X. fastidiosa medium or XFM) containing a single carbon source and no preformed amino acids has emerged as a method with a particularly high potential for the grown of X. fastidiosa and to produce genuine natural products. In this work, we identified a range of diketopiperazines from X. fastidiosa 9a5c growth in XFM, using Ultra-Fast Liquid Chromatography coupled with mass spectrometry. Diketopiperazines are reported for the first time from X. fastidiosa , which is responsible for citrus variegated chlorosis. We also report here fatty acids from X. fastidiosa , which were not biologically active as diffusible signals, and the role of diketopiperazines in signal transduction still remains unknown., Competing Interests: The authors declare no conflict of interest.

Bioassay-guided isolation of the secondary metabolites from the fungus Dichotomomyces sp . L-8 associated with the soft coral Lobophytum crassum led to the discovery of two new compounds, dichotones A and B ( 1 and 2 ), together with four known compounds including dichotocejpin C ( 3 ), bis- N -norgliovictin ( 4 ), bassiatin ( 5 ) and (3 R ,6 R )-bassiatin ( 6 ). The structures of these compounds were determined by 1D, 2D NMR and mass spectrometry. (3 R ,6 R ) - bassiatin ( 6 ) displayed significant cytotoxic activities against the human breast cancer cell line MDA-MB-435 and the human lung cancer cell line Calu3 with IC 50 values of 7.34 ± 0.20 and 14.54 ± 0.01 μM, respectively, while bassiatin ( 5 ), the diastereomer of compound 6 , was not cytotoxic.

Actinobacteria are well known for their potential in biotechnology and their production of metabolites of interest. Lichens are a promising source of new bacterial strains, especially Actinobacteria, which afford a broad chemical diversity. In this context, the culture medium of the actinobacterium Nocardia ignorata, isolated from the terrestrial lichen Collema auriforme , was studied. The strain was cultivated in a BioFlo 115 bioreactor, and the culture medium was extracted using an XAD7HP resin. Five known diketopiperazines: cyclo (l-Pro-l-OMet) ( 1 ), cyclo (l-Pro-l-Tyr) ( 2 ), cyclo (d-Pro-l-Tyr) ( 3 ), cyclo (l-Pro-l-Val) ( 4 ), cyclo (l-Pro-l-Leu) ( 5 ), and one auxin derivative: indole-carboxaldehyde ( 8 ) were isolated, along with two new brominated diketopiperazines: cyclo (d-Pro-l-Br-Tyr) ( 6 ) and cyclo (l-Pro-l-Br-Tyr) ( 7 ). Structure elucidation was performed using HRMS and 1D and 2D NMR analysis, and the synthesis of compounds 6 and 7 was carried out in order to confirm their structure.

As a part of our ongoing effort to allocate marine microbial bioactive leads, a tunicate-derived actinomycete, Streptomyces sp. Did-27, was investigated. Three new 2(1H)-pyrazinones derivatives, (S)-6-(sec-butyl)-3-isopropylpyrazin-2(1H)-one (1), (S)-3-(sec-butyl)-6-isopropylpyrazin-2(1H)-one (2) and (S)-6-(sec-butyl)-3-isobutylpyrazin-2(1H)-one (3), together with the known (1H)-pyrazinones analogues deoxymutaaspergillic acid (4), 3,6-diisobutyl-2(1H)-pyrazinone (5) and 3,6-di-sec-butyl-2(1H)-pyrazinone (6), and the diketopiperazine alkaloids cyclo(6-OH-d-Pro-l-Phe) (7), bacillusamide B (8), cyclo(l-Pro-l-Leu) and cyclo(l-Pro-l-Ile) (10) were isolated from this strain. The structures of the compounds were determined by study of their one- and two-dimensional NMR spectra as well as high-resolution mass spectral determinations. Compound 4 was reported previously as a synthetic product, while compound 6 was reported as 2-hydroxy-3,6-di-sec-butylpyrazine. Herein, we report the complete NMR data for compounds 4 and 6. The compounds were evaluated for their cytotoxic activities against three cell lines. Compound 5 showed potent and selective activity against HCT-116 cell line with IC50 of 1.5 μg/mL, while 1-10 showed variable cytotoxic activities against these cancer cell lines. These results provide further understanding about the chemistry and bioactivities of the alkylated 2(1H)-pyrazinone derivatives.

Multidrug resistance (MDR) to anticancer drugs is a serious health problem that in many cases leads to cancer treatment failure. The ATP binding cassette (ABC) transporter P-glycoprotein (P-gp), which leads to premature efflux of drugs from cancer cells, is often responsible for MDR. On the other hand, a strategy to search for modulators from natural products to overcome MDR had been in place during the last decades. However, Nature limits the amount of some natural products, which has led to the development of synthetic strategies to increase their availability. This review summarizes the research findings on marine natural products and derivatives, mainly alkaloids, polyoxygenated sterols, polyketides, terpenoids, diketopiperazines, and peptides, with P-gp inhibitory activity highlighting the established structure-activity relationships. The synthetic pathways for the total synthesis of the most promising members and analogs are also presented. It is expected that the data gathered during the last decades concerning their synthesis and MDR-inhibiting activities will help medicinal chemists develop potential drug candidates using marine natural products as models which can deliver new ABC transporter inhibitor scaffolds.

The coupling of haloarenes to styrenes and 1,1-diarylethenes has been achieved with potassium tert-butoxide in the presence of N,N'-dialkyldiketopiperazines. In contrast to previously reported reactions where phenanthroline has been used to mediate the reactions, the use of diketopiperazines can lead to either 1,1,2-triarylethenes or 1,1,2-triarylethanes, depending on the conditions used.

A new prenylated indole diketopiperazine alkaloid, cristatumin F (1), and four known metabolites, echinulin (2), dehydroechinulin (3), neoechinulin A (4) and variecolorin O (5), were isolated from the crude extract of the fungus Eurotium cristatum. The structure of 1 was elucidated primarily by NMR and MS methods. The absolute configuration of 1 was assigned using Marfey's method applied to its acid hydrolyzate. Cristatumin F (1) showed modest radical scavenging activity against DPPH radicals, and exhibited marginal attenuation of 3T3L1 pre-adipocytes.

Aqueous in situ one-pot N-Boc-deprotection-cyclization of N alpha-Boc-dipeptidyl-tert-butyl and methyl esters under microwave irradiation afforded 2,5-diketopiperazines (DKPs) in excellent yields. This protocol is rapid, safe, environmentally friendly, and highly efficient, and showed that the tert-butoxy moiety is also an excellent leaving group for these cyclizations.