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Fractionation of cytotoxic extracts of specimens of a newly described sponge genus, Candidaspongia, has yielded the candidaspongiolides (3), a complex mixture of acyl esters of a macrolide related to tedanolide. The general structure of the candidaspongiolides was determined by analyses of various 2D NMR and MS data sets. The acyl ester componentswere identified by GC-MS analysis of the derived fatty acid methyl esters. The mixture could be selectively converted to the deacylated macrolide core (4) by enzymolysis with immobilized porcine lipase, with the structure of the candidaspongiolide core then secured by NMR and MS analysis. The candidaspongiolide mixture was potently cytotoxic, exhibiting a mean panel 50% growth inhibition (GI50) of 14 ng/mL in the National Cancer Institute’s 60-cell-line in Vitro antitumor screen.

The isolation of taxol from Taxus x. media Hicksii includes four taxinines which co-elute and interfere with HPLC analyses of taxol. Their structures: 1,9α-dihydroxy-2α,10β-diacetoxy-5α-cinnamatetaxa-4(20),11-diene-13-one; 1,10β-dihydroxy-2α,9α-diacetoxy-5α-cinnamatetaxa-4(20),11-diene-13-one;2α,9α-dihydroxy-10β,13α-diacetoxy-5α-cinnamatetaxa-4(20),11-diene and; 2α,10β-dihydroxy-9α,13α-diacetoxy-5α-cinnamatetaxa-4(20),11-diene, have been determined by 1D(1H, 13C) and 2D(DQCOSY, TOCSY, HMQC, HMBC and TROESY) NMR spectroscopy-Transacetylation between 10β and 9α on the baccatin core has been documented to occur in CDCl3 and CD3OD at 27°. Interatomic distances were obtained from TROESY build-up curves of the compound and were compare to those obtained from a computer minimized structure.

Aqueous extracts of many marine invertebrates have exhibited some activity in the National Cancer Institute's primary screen for anti-HIV cytopathicity. Using a variety of techniques, including gel permeation, size exclusion and ion exchange chromatography, toluidine blue metachromicity,13C-NMR spectroscopy and combustion analyses, we have determined that this activity is largely due to sulphated polysaccharides. Because of the wide occurrence of this class of compounds in these organisms we sought a method for the rapid dereplication of sulphated polysaccharides. It was critical that the method selected for dereplication allow differentiation of anionic polysaccharides from other AIDS-antiviral chemotypes. After evaluating a variety of methods, we found that the most efficient strategy appeared to be precipitation of the polysaccharide fraction from aqueous ethanolic solutions of the crude aqueous extracts.

Lipophilic and hydrophilic extracts from approximately 600 strains of cultured mcyanophytes, representing some 300 species, were examined for antiviral activity against three pathogenic viruses. Approximately 10% of the cultures produced substances that caused significant reduction in cytopathic effect normally associated with viral infection. The screening program identified the order Chroococcales as commonly producing antiviral agnets.

The novel sulfated sterol ibisterol sulfate ( 1 ) was isolated from the deep water Caribbean sponge Topsentia sp. The combination of both a Δ 9(11) olefin and a methyl group at C-14 has not previously been reported in sponge sterols.

A polyhalogenated acyclic monoterpene, 6(R)-bromo-3(S)-(bromomethyl)-7- methyl-2,3,7-trichloro-1-octene (1) was obtained as a major component of the organic extract of the red alga Portieria hornemannii. X-ray diffraction analysis provided the complete structure, including correct placement of the different halogen atoms and determination of the absolute stereochemistry. Detailed NMR analyses provided complete 1H and 13C assignments. Compound 1 exhibited highly differential cytotoxicity against the U.S. National Cancer Institute's new in vitro human tumor cell line screening panel; brain tumor, renal, and colon tumor cell lines were most sensitive to 1, while leukemia and melanoma lines were relatively less sensitive. A second collection of P. hornemanni yielded the novel, monocyclic 2, considerably less cytotoxic and devoid of differential activity. On the basis of its unprecedented cytotoxicity profile in the NCI primary screen, compound 1 has been selected by the NCI Decision Network Committee for preclinical drug development.

Between 1960 and 1981 the National Cancer Institute (NCI) screened 114,000 extracts of 35,000 plants, mainly collected in temperate regions. Of the three clinically active anticancer drugs so far discovered in that programme, none was isolated from a plant collected on an ethnobotanical basis, though various Taxus species, which are the source of taxol, are reported to have been used medicinally. Since 1986, the NCI has focused its collections in tropical and subtropical regions worldwide; collections cover a broad taxonomic range, though priority is given to medicinal plants when relevant information is available. As of August 1993, 21,881 extracts derived from over 10,500 samples had been tested in a screen for activity against the human immunodeficiency virus (HIV); 2320 of these extracts were of medicinal plant origin. Approximately 18% of both the total number of extracts and the medicinal plant-derived extracts showed significant anti-HIV activity; in each instance about 90% of the active extracts were aqueous. The activity of the aqueous extracts has been attributed mainly to the presence of polysaccharides or tannins. Four plant-derived compounds are in preclinical development at the NCI; only one of the four sources plants, obtained from a noncontract source, was collected on an ethnobotanical basis. At this stage the results indicate that the current NCI collection policy offers the best chances for the discovery and development of agents for the treatment of AIDS (acquired immune deficiency syndrome) and cancer.

This review is an updated and expanded version of a paper that was published in this journal in 1997. The time frame has been extended in both directions to include the 22 years from 1981 to 2002, and a new secondary subdivision related to the natural product source but applied to formally synthetic compounds has been introduced, using the concept of a "natural product mimic" or "NM" to join the original primary divisions. From the data presented, the utility of natural products as sources of novel structures, but not necessarily the final drug entity, is still alive and well. Thus, in the area of cancer, the percentage of small molecule, new chemical entities that are nonsynthetic has remained at 62% averaged over the whole time frame. In other areas, the influence of natural product structures is quite marked, particularly in the antihypertensive area, where of the 74 formally synthetic drugs, 48 can be traced to natural product structures/mimics. Similarly, with the 10 antimigraine drugs, seven are based on the serotonin molecule or derivatives thereof. Finally, although combinatorial techniques have succeeded as methods of optimizing structures and have, in fact, been used in the optimization of a number of recently approved agents, we have not been able to identify a de novo combinatorial compound approved as a drug in this time frame.

No abstract available

Micellar electrokinetic chromatography (MEKC) was applied to the separation of the anti-HIV agents, michellamines A and B, and two other structurally related monomers found in the extract of the Ancistrocladus plants. Using buffers containing either 10 mM sodium phosphate (pH 7.0), 50 mM sodium deoxycholate and 10-20% acetonitrile or 5 mM sodium phosphate (pH 7.0), 20 mM sodium dodecyl sulfate and 25% acetonitrile allowed baseline separations of the four components in the mixture in less than 10 min. The MEKC methods gave sharper peaks and better resolution compared to high-performance liquid chromatography. For MEKC separation of the plant extracts, UV absorption detection provided adequate sensitivity; however, higher sensitivity could be achieved with UV laser-induced fluorescence detection (LIF). Using the sodium dodecyl sulfate-containing buffer and LIF, the limit of detection for michellamine B was approximately 2 ng/mL. The sensitivity was degraded approximately 100-fold when using the deoxycholate buffer because of high background fluorescence. Preliminary results show that MEKC with LIF is feasible for the sensitive detection of michellamine B in serum.

High-performance liquid chromatography (HPLC) and micellar electrokinetic chromatography (MEKC) were applied for the separation of taxol, cephalomannine, and baccatin III in crude extracts from the needle and bark of Taxus species. The chromatogram of the bark extract was cleaner than that of the needle allowing a more reliable detection of taxol and cephalomannine in the bark extract. However, HPLC quantitation of taxol in the needle extract would be difficult due to coeluting taxinines. Nevertheless, this was not a problem in the MEKC experiment. In comparison to HPLC, MEKC offered baseline resolution of taxol from taxinines in the needle extract, less solvent waste, a smaller sample requirement, and the simultaneous detection of taxol, cephalomannine and baccatin III in a relatively simpler electrophoretic run.

A novel process was designed for the large-scale isolation of bryostatin 1 from the bryozoan Bugula neritina L. in order to obtain multigram quantities of highly pure material for formulation studies, preclinical toxicology, and clinical trials in cancer patients. Multigram quantities of bryostatin 1 were obtained from a collection of approximately 10,000 gallons of wet animal. A phorbol dibutyrate (PDBu) receptor binding assay and hplc with photodiode array detection were used for the design, validation, and control of the isolation process.

1-Vinyl-8-hydroxy-beta-carboline was identified as the cytotoxic constituent of the bryozoans Catenicella cribraria and Cribricellina cribraria. Literature nmr data for this previously known compound, now reported from a new source, were corrected.

Condensation of 3,5-diacylpyrantriones with various aromatic amines gave a new class of potent, orally active, antiallergic compounds, the 3-[(arylamino)ethylidene]-5-acylpyrantriones, hereafter referred to as pyranenamines, as evaluated not only in the traditional rat passive cutaneous anaphylaxis (PCA) assay but also in the in vitro fragmented rat and primate lung assay. Potencies in the PCA system, when measured intravenously, reached a maximum ID50 of 0.9 mu/kg (1000 times more potent than disodium chromoglycate) with 5-acetyl-4-hydroxy-3-[1-[(3,5-bis-glyceramoylphenyl)amino]ethylidene]-2H-pyran-2,6(3H)-dione (100), as predicted by structure-activity relationship (SAR) analysis. Potencies in the iv PCA system correlated well with potencies in the in vitro rat lung system but not with potencies in the oral PCA system or the in vitro primate lung system. Several compounds had good oral potency, and one analogue, 3-acetyl-4-hydroxy-3-[1-[3-amino-4-hydroxyphenyl)amino]ethylidene]-2H-pyran-2,6(3H)-dione hydrochloride (78), reached an oral ID50 of less than 1 mg/kg and was better than 10 times more effective than disodium chromoglycate at inhibiting the release of histamine and slow-reacting substance of anaphylaxis in the fragmented primate lung assay.

Duryne, 1 , a cytotoxic metabolite which inhibits the growth of both mouse and human tumor cell lines in vitro has been isolated from the marine sponge Cribrochalina dura . Its structure has been assigned by spectral methods.

A new glycopeptide antibiotic complex was isolated from the fermentation culture of Kibdelosporangium aridum subsp. largum (SKF AAD-609) by affinity chromatography on a D-alanyl-D-alanine agarose column. This major components of the complex were resolved by preparative reversed-phase HPLC. Mild acid hydrolysis showed that the new antibiotics have the same mannosyl aglycon (2) as the aridicins. FAB mass spectrometry, isoelectric focusing, potentiometric titration and carbohydrate and fatty acid analyses were used to determine the structures of the five major components of the complex. These studies showed that the kibdelins differ from the aridicins only in the oxidation level at the C-6 position of the amino sugar. Kibdelin A (5), B (6), C1 (7), C2 (8) and D (9) are a series of N-acylglucosamine analogs containing saturated straight and branched chain C10-C12 fatty acids whereas, in kibdelin D the fatty acid component is (Z)-4-decenoic acid.

Chlorocardicin, a novel monocyclic beta-lactam, was isolated from the fermentation broth of a Streptomyces sp. by the use of non-ionic porous resin and reverse phase chromatography. This chlorine-containing antibiotic is structurally related to nocardicin A. Its physico-chemical characteristics and detailed NMR analysis are described.

A recently developed tetrazolium-based microculture assay was used to screen extracts of cultured cyanobacteria (blue-green algae) for inhibition of the cytopathic effects of the human immunodeficiency virus (HIV-1), which is implicated as a causative agent of AIDS. A number of extracts were found to be remarkably active against the AIDS virus. A new class of HIV-1-inhibitory compounds, the sulfonic acid-containing glycolipids, was discovered through the use of the microculture assay to guide the fractionation and purification process. The pure compounds were active against HIV-1 in cultured human lymphoblastoid CEM, MT-2, LDV-7, and C3-44 cell lines in the tetrazolium assay as well as in p24 viral protein and syncytium formation assays.

The ketone group in 4-keto-3,4-dihydrosulfostyril (I) displays anomalous properties. Its reactions are described; in some reactions it behaved normally, in others it proved quite inert. Attempted oximination of I led to extrusion of the sulfone group with formation of isatin derivatives. Compound I was converted to a tetracyclic indole derivative.

A new antibacterial antibiotic complex, aridicin, was produced by a new genus, Kibdelosporangium aridum (SK&F-AAD-216). The individual factors, aridicins A, B and C, were isolated from the fermentation broth by an Amberlite XAD-7 resin extraction and purified by preparative reversed phase HPLC. The aridicins were found to be novel members of the glycopeptide class of antibiotics as exemplified by ristocetin and vancomycin, based on chemical and spectroscopic data, their molecular weights as determined by FAB mass spectrometry (1,786, 1,800 and 1,814), the detection of actinoidinic acid in their acid hydrolysates, and detailed TLC and HPLC comparisons with representative members of this class.

A set of four cerebrosides was isolated from a Pachybasium species and purified by preparative reversed-phase HPLC. All four products displayed activity in a natural product screen aimed at detecting novel cell wall-active antifungal agents based on synergy with the known glucan synthetase inhibitor, aculeacin. Based on degradation studies, fast atom bombardment mass spectrometry and 13C and high field 1H NMR techniques, the structure of the major cerebroside was determined to be (4E,8E)-N-D-2'-hydroxy-(E)-3'- hexadecenoyl-1-O-beta-D-glucopyranosyl-9-methyl-4,8-sphingadiene. The other components were found to be the corresponding 2'-hydroxypalmitic acid analog with one less double bond and an analogous pair containing 2'-hydroxystearic acid with and without the 3' double bond.

The aglycone and two pseudoaglycones of aridicin A were prepared by selective hydrolysis and characterized, chemically and biologically. These new analogs demonstrate improved activities in vitro over the parent antibiotics against methicillin sensitive and resistant staphylococci. The major determinant of activity is the mannose substituent, the presence of which results in less potent compounds. The analogs have potent activity against enterococci.

QSAR techniques played a major role in development of the antiallergic pyranenamines (I). Graphical analysis of data resulting from an unsuccessful Topliss approach suggested that increased substituent hydrophilicity might enhance potency. The 3-NHAc-4-OH derivative which first resulted was an order of magnitude more potent than any preceding series member, and its deacylated congenar is clinical candidate SK&F 78729 (R1 = -NH2, R2 - OH, R3 = H). Further pursuit of hydrophilicity and other strategies suggested by multiple regression yielded 98 pyranenamines, the most active [R1 = R3 = NHCO(CHOH)2H, R2 = H] being 1000 times more potent than any original series member.

An affinity support was designed to facilitate the isolation and purification of glycopeptide antibiotics by mimicking their known affinity for the bacterial cell wall. Members of this class of antibiotics inhibit peptidoglycan biosynthesis by specifically binding to pentapeptide precursors terminating with L-Lys-D-Ala-D-Ala. A series of ligands (Gly, D-Ala, D-Ala-D-Ala and alpha-N-Ac-L-Lys-D-Ala-D-Ala) were immobilized on an N-hydroxysuccinimide-activated agarose support and evaluated using the glycopeptides vancomycin and the aridicin complex. Conditions were developed to enable complete adsorption and efficient elution of both antibiotics. Of the four ligands, the readily available dipeptide offered the best compromise between high binding specificity and recovery on elution. Binding and subsequent high recovery of biologically active products were observed for eight other glycopeptide antibiotics. Column performance was shown by purification of vancomycin directly from a low titer fermentation broth. The applicability of this technique to large scale isolation was demonstrated by the preparative affinity chromatography of 36 g of the aridicins.