Pure compound libraries; a new perspective for natural product based drug discovery.

What often distinguishes the leaders in drug discovery and development from the rest is the quality of their compound libraries and the ease of access that they have to the information within those libraries. The screening of natural products can provide greater structural diversity than standard synthetic chemistry and offers significant opportunities for finding novel low molecular weight lead compounds. However, which strategy is the best for natural-product-based drug discovery? Two well established but relatively time consuming approaches are the screening of crude extracts and pre-fractionated extracts. This case study describes a third, pure-compound-screening approach, and discusses its benefits and pitfalls.

Function of glycosyltransferase genes involved in urdamycin A biosynthesis.

BACKGROUND: Urdamycin A, the principle product of Streptomyces fradiae Tü2717, is an angucycline-type antibiotic. The polyketide-derived aglycone moiety is glycosylated at two positions, but only limited information is available about glycosyltransferases involved in urdamycin biosynthesis. RESULTS: To determine the function of three glycosyltransferase genes in the urdamycin biosynthetic gene cluster, we have carried out gene inactivation and expression experiments. Inactivation of urdGT1a resulted in the predominant accumulation of urdamycin B. A mutant lacking urdGT1b and urdGT1c mainly produced compound 100-2. When urdGT1c was expressed in the urdGT1b/urdGT1c double mutant, urdamycin G and urdamycin A were detected. The mutant lacking all three genes mainly accumulated aquayamycin and urdamycinone B. Expression of urdGT1c in the triple mutant led to the formation of compound 100-1, whereas expression of urdGT1a resulted in the formation of compound 100-2. Co-expression of urdGT1b and urdGT1c resulted in the production of 12b-derhodinosyl-urdamycin A, and co-expression of urdGT1a, urdGT1b and urdGT1c resulted in the formation of urdamycin A. CONCLUSIONS: Analysis of glycosyltransferase genes of the urdamycin biosynthetic gene cluster led to an unambiguous assignment of each glycosyltransferase to a certain biosynthetic saccharide attachment step.

New trichothecenes isolated from Holarrhena floribunda.

Bioassay-guided fractionation of an extract of Holarrhena floribunda stem, has led to the isolation of the new trichothecenes, 8-dihydrotrichothecinol A (1), loukacinol A (2), and loukacinol B (3), and the known compounds, trichothecolone (4), trichothecin (5), trichothecinol A (6), rosenonolactone (7), 6beta-hydroxyrosenonolactone (8), and rosololactone (9). The structures were determined by spectral and chemical methods, and absolute configurations were established by a modified Horeau's method using HPLC. Compounds 1 and 6 exhibited significant cytotoxicity against several human tumor cell lines, whereas compound 8 showed moderate and weak antileishmanial activity toward extracellular and intracellular Leishmania donovani, respectively.

Antimicrobial, antitumor and antileishmania screening of medicinal plants from Guinea-Bissau.

Following an ethnobotanical search carried out in Guinea-Bissau, eighteen extracts derived from sixteen medicinal species were screened for antimicrobial, antitumor and antileishmania activity. Significant antitumor activity was found for Holarrhena floribunda against KB (squamous carcinoma), SK-Mel 28 (melanoma), A 549 (lung carcinoma) and MDA-MB 231 (mamma carcinoma) cell lines, with corresponding IC50 values of 7.9, 9.0, 3.4 and 9.9 micrograms/ml. Khaya senegalensis and Anthostema senegalense exhibited a significant activity against Leishmania donovani with IC50 values of 9.8 and 9.1 micrograms/ml, respectively. Most of the extracts showed week or moderate antibacterial and antifungal activity, with MIC values in the range 0.25-1.0 mg/ml. Active extracts were submitted to bioassay-guided fractionation, and the IC50 and MIC of the active fractions were determined.

Aphidicolin glycinate inhibits human neuroblastoma cell growth in vivo.

Aphidicolin is a fungal derived tetracyclic diterpene antibiotic. It is selectively toxic for neuroblastoma (NB) cells in vitro but has no significant effects on the viability of normal human cells and a variety of other tumor entities. We evaluated the antitumoral effects of the water soluble ester aphidicolin glycinate (AphiG) on established human NB xenografts from UKF-NB-3 cells in athymic (nude) mice. Furthermore, we explored the efficacy of direct intraneoplastic and systemic delivery of AphiG. Systemic administration of AphiG (60 mg/kg intraperitoneally, twice per day on 10 consecutive days) significantly suppressed tumor growth but was not able to induce any cures. In contrast, intratumoral AphiG injections (60 or 40 mg/kg/twice a day for 4 days) induced complete tumor regression. Two weeks after the end of treatment no tumor cells were microscopically detectable. Animals were free of tumor for more than 90 days. Histologic examination of inner organs and bone marrow did not reveal any apparent toxic effects of AphiG. These data strongly indicate that AphiG deserves further evaluation as a specific treatment for neuroblastoma.

Self-report ratings and informants' ratings of personalities of depressed outpatients.

OBJECTIVE: This study sought to determine whether personality traits of depressed patients could be assessed similarly by informants and self-reports of the patients themselves. METHOD: Forty-six depressed outpatients completed the self-report (first-person) version of the Revised NEO Personality Inventory and nominated informants who knew them well to complete the third-person version of that instrument. RESULTS: Agreement between the self-ratings and informants' ratings on the five factors of the inventory--neuroticism, extraversion, openness-to-experience, agreeableness, and conscientiousness--was high. The only significant difference between the self-ratings and informants' ratings was on the extraversion scale, where the patients rated themselves as significantly more introverted than did the informants. CONCLUSIONS: Informants' ratings of personality are similar to self-report ratings of depressed patients. Depressed mood may not influence the self-report of personality traits.

Relationship between the five-factor model of personality and unipolar, bipolar and schizophrenic patients.

The purpose of this study was to examine personality differences among three different Axis I disorders-recovered patients with unipolar depression (n = 62), euthymic patients with bipolar disorder (n = 34), and patients with schizophrenia in the residual phase of their illness (n = 41) using the five-factor model of personality (FFM). The dimensions of the FFM-Neuroticism (N), Extraversion (E), Openness (O), Agreeableness (A), and Conscientiousness (C)-were measured with composite scores derived from the NEO Personality Inventory (NEO PI) and the Revised NEO Personality Inventory (NEO PI-R). While no group differences emerged on N or C, the bipolar patients scored significantly higher on the Positive Emotion facet (subscale) of E than the unipolar patients. The schizophrenic patients scored lower on the Feelings, Values and Actions facets of O than did the unipolar and bipolar patients. The unipolar patients scored higher on A than the schizophrenic patients.

Cloning and insertional inactivation of Streptomyces argillaceus genes involved in the earliest steps of biosynthesis of the sugar moieties of the antitumor polyketide mithramycin.

Two genes (mtmD and mtmE) were cloned and sequenced from the mithramycin producer Streptomyces argillaceus. Comparison with proteins in databases and enzymatic assays after expression in Escherichia coli showed that they encode a glucose-1-phosphate:TTP thymidylyl transferase and a TDP-D-glucose 4,6-dehydratase, respectively. The mtmD gene was inactivated by gene replacement, generating a nonproducing mutant that accumulates a tetracyclic compound designated premithramycinone. The identification of premithramycinone reveals new aspects of the mithramycin biosynthetic pathway and suggests that at least some glycosylations occur before breakage of the fourth ring.

Bipolar disorder, unipolar depression and the Five-Factor Model of personality.

We examined differences between personality characteristics of euthymic bipolar disorder patients (BD) (n = 34) and recovered unipolar depressed patients (UD) (n = 74) using the taxonomy of the Five-Factor Model of personality (FFM) as measured by composite scales derived from the NEO Personality Inventory (NEO PI) and the revised NEO PI (NEO PI-R). Euthymic BD patients scored significantly higher on the Openness (O) dimension and the Positive Emotions facet of the E dimension than did recovered UD patients. For O, euthymic BD patients scored higher on the Feelings facet. These results suggest not only that euthymic BD patients are more likely to experience positive affects than recovered UD patients, but also that euthymic BD patients are more receptive to their positive and negative feelings than are recovered UD patients.

Balmoralmycin, a new angucyclinone, and two related biosynthetic shunt products containing a novel ring system.

A new angucyclinone, named balmoralmycin (1), was isolated as an inhibitor of protein kinase C-alpha (PKC-alpha) from the Streptomyces strain P6417. Chemical screening of extracts of the same strain resulted in the detection of two decaketides with unusual structural features (2 and 3). Both compounds belong to a recently described structural class of secondary metabolites which arises from engineered biosynthesis of a recombinant Streptomyces strain. The isolation of compounds of this class from a wild-type strain has never been reported before.

Inhibitory effect of modified bafilomycins and concanamycins on P- and V-type adenosinetriphosphatases.

Various ATPases have been tested for their sensitivity to naturally occurring unusual macrolides and their chemically modified derivatives, which are structurally related to bafilomycin A1 (1), the first specific inhibitor of vacuolar ATPases. The structure-activity study showed that in general the concanamycins, 18-membered macrolides, are better and more specific inhibitors than the bafilomycins of this class of membrane-bound ATPases. The additional carbohydrate residue is not responsible for the improved activity. The importance of an intact hemiketal ring, which is part of an intramolecular hydrogen-bonding network, and the effects of the size of the macrolactone ring are discussed. The structurally related elaiophylin (13), a C2-symmetric macrodiolide antibiotic, proved to be inactive on vacuolar ATPases but still retained its inhibitory effect on P-type ATPases.