Miniaturized Cultivation of Microbiota for Antimalarial Drug Discovery.

The ongoing search for effective antiplasmodial agents remains essential in the fight against malaria worldwide. Emerging parasitic drug resistance places an urgent need to explore chemotherapies with novel structures and mechanisms of action. Natural products have historically provided effective antimalarial drug scaffolds. In an effort to search nature's chemical potential for antiplasmodial agents, unconventionally sourced organisms coupled with innovative cultivation techniques were utilized. Approximately 60,000 niche microbes from various habitats (slow-growing terrestrial fungi, Antarctic microbes, and mangrove endophytes) were cultivated on a small-scale, extracted, and used in high-throughput screening to determine antimalarial activity. About 1% of crude extracts were considered active and 6% partially active (≥ 67% inhibition at 5 and 50 µg/mL, respectively). Active extracts (685) were cultivated on a large-scale, fractionated, and screened for both antimalarial activity and cytotoxicity. High interest fractions (397) with an IC50 < 1.11 µg/mL were identified and subjected to chromatographic separation for compound characterization and dereplication. Identifying active compounds with nanomolar antimalarial activity coupled with a selectivity index tenfold higher was accomplished with two of the 52 compounds isolated. This microscale, high-throughput screening project for antiplasmodial agents is discussed in the context of current natural product drug discovery efforts.

Screening mangrove endophytic fungi for antimalarial natural products.

We conducted a screening campaign to investigate fungi as a source for new antimalarial compounds. A subset of our fungal collection comprising Chinese mangrove endophytes provided over 5000 lipophilic extracts. We developed an accelerated discovery program based on small-scale cultivation for crude extract screening and a high-throughput malaria assay. Criteria for hits were developed and high priority hits were subjected to scale-up cultivation. Extracts from large scale cultivation were fractionated and these fractions subjected to both in vitro malaria and cytotoxicity screening. Criteria for advancing fractions to purification were developed, including the introduction of a selectivity index and by dereplication of known metabolites. From the Chinese mangrove endophytes, four new compounds (14-16, 18) were isolated including a new dimeric tetrahydroxanthone, dicerandrol D (14), which was found to display the most favorable bioactivity profile.

CNS and antimalarial activity of synthetic meridianin and psammopemmin analogs.

The marine invertebrate-derived meridianin A, the originally proposed structure for psammopemmin A, and several related 3-pyrimidylindole analogs were synthesized and subsequently investigated for central nervous system, antimalarial, and cytotoxic activity. A Suzuki coupling of an indoleborate ester to the pyrimidine electrophile was utilized to form the natural product and derivatives thereof. The 3-pyrimidineindoles were found to prevent radioligand binding to several CNS receptors and transporters, most notably, serotonin receptors (<0.2 µM K(i) for 5HT(2B)). Two compounds also inhibited the human malaria parasite Plasmodium falciparum (IC(50) <50 µM). Only the natural product was cytotoxic toward A549 cells (IC(50)=15 µM).