There is increasing evidence that uncultivated bacterial symbionts are the true producers of numerous bioactive compounds isolated from marine sponges. The localization and heterologous expression of biosynthetic genes could clarify this issue and provide sustainable supplies for a wide range of pharmaceuticals. However, identification of genes in the usually highly complex symbiont communities remains a challenging task. For polyketides, one of the most important groups of sponge-derived drug candidates, we have developed a general strategy that allows one to rapidly access biosynthetic gene clusters based on chemical moieties. Using this method, we targeted polyketide synthase genes from two different sponge metagenomes. We have obtained from a sponge-bacterial association a complete pathway for the rare and potent antitumor agent psymberin from Psammocinia aff. bulbosa. The data support the symbiont hypothesis and provide insights into natural product evolution in previously inaccessible bacteria.

The present study examined the role of xanthohumol (XN), a plant chalcone, on apolipoprotein B (apoB) and triglyceride (TG) synthesis and secretion, using HepG2 cells as the model system. The data indicated that XN decreased apoB secretion in a dose-dependent manner under both basal and lipid-rich conditions (as much as 43% at 15 mol/L). This decrease was associated with increased cellular apoB degradation. To determine the mechanism underlying this effect, we examined triglyceride availability, a major factor in the regulation of apoB secretion. XN inhibited the synthesis of TG in the microsomal membrane and the transfer of this newly synthesized TG to the microsomal lumen (decreases of 26 and 64%, respectively, under lipid-rich conditions), indicating that TG availability is a determining factor in the regulation of apoB secretion under the experimental conditions. The inhibition of TG synthesis was caused by a reduction in diacylglycerol acyltransferase (DGAT) activity, which corresponded to a decrease in DGAT-1 mRNA expression, but not DGAT-2 expression. Microsomal triglyceride transfer protein (MTP) may also control the rate of TG transfer from the microsomal membrane to the active lumenal pool. XN decreased MTP activity in a dose-dependent manner (as much as 30%). Whether the reduction in TG accumulation in the microsomal lumen is predominantly due to DGAT and/or MTP activity remains unknown. In summary, the data suggest that xanthohumol is a potent inhibitor of apoB secretion.

Human African trypanosomiasis (HAT), also known as African sleeping sickness, is a neglected tropical disease with inadequate therapeutic options. We have launched a collaborative new lead discovery venture using our repository of extracts and natural product compounds as input into our growth inhibition primary screen against Trypanosoma brucei. Careful evaluation of the spectral data of the natural products and derivatives allowed for the elucidation of the absolute configuration (using the modified Mosher's method) of two new peroxiterpenes: (+)-muqubilone B (1a) and (−)-ent-muqubilone (3a). Five known compounds were also isolated: (+)-sigmosceptrellin A (4a), (+)-sigmosceptrellin A methyl ester (4b), (−)-sigmosceptrellin B (5), (+)-epi-muqubillin A (6) and (−)-epi-nuapapuin B methyl ester (7). The isolated peroxiterpenes demonstrated activities in the range from IC 50 = 0.2 -2 μg/mL.The search for chemotherapeutics to treat human African trypanosomiasis (HAT), also known as African sleeping sickness, has not been a major target of industry drug discovery campaigns, even though the currently available therapeutics are inadequate.1 This is surprising as there are 50,000 annual cases of infection, HAT is the world's third most devastating parasitic disease, and it remains a major threat to more than 60 million people.2 -4 Because it is a serious health problem for resource poor regions of Africa, HAT is designated as a neglected tropical disease (NTD). The causative agent is a protozoan parasite, Trypanosoma brucei, 5 subdivided in two sub-genera, T. brucei gambiense in West Africa and T. brucei rhodesiense in East Africa. 1 Most of the four current chemotherapeutics to combat these parasites at their different development stages are very antiquated. The drugs (by registration date for HAT treatment) consist of suramin4 (1922), pentamidine1 (1941), melarsoprol3 (1949), and eflorithine1 (1990). Another relevant point is that new strains of T. brucei are showing cross-resistance to some of these agents.6 In recent years, only one compound, DB289 7 (a synthetic analog of pentamidine), received serious clinical evaluation against HAT, but further development was discontinued in 2008. 3 * To whom correspondence should be addressed. phil@chemistry.ucsc.edu. Tel: (831)

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Bouillomides A (1) and B (2) are two depsipeptide analogues of dolastatin 13. Isolated from a Guamanian sample of Lyngbya bouillonii, the planar structures were elucidated on the basis of HR-ESI-MS and NMR data, while the absolute configurations were determined by employing functional group conversions, modified Marfey’s analysis, and detailed analyses of ROESY correlations. Compounds 1 and 2 selectively inhibited serine proteases elastase (IC50 = 1.9 μM for both) and chymotrypsin (IC50 = 0.17 and 9.3 μM, respectively) while showing no inhibition of trypsin (IC50 > 100 μM).

A new meroditerpene, (+)-isojaspic acid (1), along with two known meroditerpenes, cacospongin D (2) and jaspaquinol (3), have been isolated from a marine sponge Cacospongia. Comprehensive taxonomic identification distinguished this Cacospongia apart from morphologically similar Psammocinia. The absolute stereochemistry of 1 was elucidated on the basis of extensive 1D and 2D NMR techniques and analysis of the optical rotation versus (+)-zonarol (8), (+)-isozonarol (9), (-)-dactylosponol (10), and (+)-hyatellaquinone (11). Furthermore, bioactivity evaluation showed that the meroditerpenes isolated significantly inhibited Staphylococcus epidermidis.

The dramatic biogeographical variations in the secondary metabolites from Psammocinia aff. bulbosa have complicated our efforts to re-isolate the two most cytotoxic of its metabolites, (+)-psymberin and (+)-cyclocinamide A. Reported now are the results of a new study that demonstrates our ability to repeatedly isolate these two compounds through targeted collection efforts. Additional study of the new sample of (+)-cyclocinamide A has enabled finalizing its biological activity and absolute stereochemistry as 4S, 7S, 11S, 14S.In the early 1990s we began a campaign to survey the chemical ecology of Psammocinia aff. bulbosa communities from Papua New Guinea. It is now clear that a side-by-side chemical and taxonomic examination is obligatory to differentiate among populations of Psammocinia (Dictyoceratida, Irciniidae), Cacospongia (Dictyoceratida, Thorectidae), or Ircinia (Dictyoceratida, Irciniidae) 1 having very similar morphologies. Recognizing this critical step made it possible to engage in a meaningful synopsis of the chemical variations observed for a small library of P. aff. bulbosa specimens. Reported now are the final results of a project whose goals were to: (a) obtain fresh sponge specimens containing both psymberin and cyclocinamide A in order to facilitate on-going molecular genetics studies on their biosynthetic pathways, 2 (b) complete the full absolute chirality assignment for cyclocinamide A, 3,4 and (c) accumulate additional biological activity data for cyclocinamide A. 3 Outlined in Table 1 are an astonishing variety of biosynthetic products that we have observed to date from six distinct collections whose voucher material has each been identified as Psammocinia aff. bulbosa. The major constituents of specimens I -V range from sesterterpenes (variabilin 5 ), polyketides (preswinholide A 6 and swinholide A 6,7 ), a cyano-sponge cyclic hexapeptide (psymbamide A 7 ), a halogenated hexapeptide (cyclocinamide A 3 ), to a NRPS-PKS mixed biogenetic derived compound (psymberin 8 ). By contrast, meroterpenes (cacospongin B 9 and chromarol D 9 ) usually observed from sponges of Cacospongia-genus were isolated from sample VI. Though samples V and VI were obtained from adjacent sites, the chemical profiles of their major constituents clearly showed the biosynthetic machinery of these two sponge collections was distinct. These plus the other dramatic biogeographical *To whom correspondence should be addressed. phil@chemistry.ucsc.edu, Tel: (831) 459-2603. Fax: (831) Table 1 complicated our efforts to re-isolate the two most structurally unique of these metabolites, psymberin and cyclocinamide A.Before proceeding, it is important to briefly review the historical record highlighted in Figure 1 pertaining to absolute stereochemical assignments for psymberin and cyclocinamide A. The 5S, 8S, 9S, 11R, 13R, 15S, 16R, 17R structure we originally proposed for (+)-psymberin 8 (1a) was reaffirmed from two independent total syntheses of 1c. 10,11 The synthetic products also provided the basis for the 4S ...

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