High In Vitro and In Vivo Activity of BI-847325, a Dual MEK/Aurora Kinase Inhibitor, in Human Solid and Hematologic Cancer Models.

BI-847325 is an ATP-competitive inhibitor of MEK/Aurora kinases with the potential to treat a wide range of cancers. In a panel of 294 human tumor cell lines in vitro, BI-847325 was found to be a highly selective inhibitor that was active in the submicromolar range. The most sensitive cancer types were acute lymphocytic and myelocytic leukemia, melanomas, bladder, colorectal, and mammary cancers. BI-847325 showed a broader range of activity than the MEK inhibitor GDC-0623. The high efficacy of BI-847325 was associated with but not limited to cell lines with oncogenic mutations in NRAS, BRAF, and MAP2K1.The high antiproliferative activity of BI-847325 was validated in vivo using subcutaneous xenograft models. After oral administration of 80 and 40 mg/kg once weekly for 3 or 4 weeks, BI-847325 was highly active in four of five colorectal, two of two gastric, two of two mammary, and one of one pancreatic cancer models (test/control < 25%), and tumor regressions were observed in five of 11 cancer models. The treatment was well tolerated with no relevant lethality or body weight changes. In combination with capecitabine, BI-847325 displayed synergism over single-agent therapies, leading to complete remission in the triple-negative mammary model MAXFTN 401, partial regression in the colon model CXF 1103, and stasis in the gastric models GXA 3011 and GXA 3023. In conclusion, dual MEK/Aurora kinase inhibition shows remarkable potential for treating multiple types of hematologic and solid tumors. The combination with capecitabine was synergistic in colorectal, gastric, and mammary cancer. SIGNIFICANCE: We report the preclinical evaluation of BI-847325, a MEK/Aurora kinase inhibitor. Our data demonstrate that BI-847325 has potent antitumor activity in a broad range of human solid and hematologic cancer models in vitro and in vivo and is well tolerated in animal models. It also shows synergistic effect when combined with capecitabine. These findings provide a strong rationale for further development of BI-847325 as a potential therapeutic for patients with cancer.

Mansouramycins E-G, Cytotoxic Isoquinolinequinones from Marine Streptomycetes.

Chemical investigation of the ethyl acetate extract from the marine-derived Streptomyces sp. isolate B1848 resulted in three new isoquinolinequinone derivatives, the mansouramycins E-G (1a-3a), in addition to the previously reported mansouramycins A (5) and D (6). Their structures were elucidated by computer-assisted interpretation of 1D and 2D NMR spectra, high-resolution mass spectrometry, and by comparison with related compounds. Cytotoxicity profiling of the mansouramycins in a panel of up to 36 tumor cell lines indicated a significant cytotoxicity and good tumor selectivity for mansouramycin F (2a), while the activity profile of E (1a) was less attractive.

Boshramycinones A-C: New anthracyclinones produced by a marine-derived Streptomyces sp.: isolation, structure elucidation and biological activities.

Boshramycinones A-C (1-3), three new anthracyclinones, were isolated from the culture broth of the marine-derived Streptomyces sp. Mei 16-1,2 together with 2-acetyl-1,8-dihydroxy-3-methyl-anthraquinone (4) and bafilomycins B1, B2, and C1-amide. The isolated compounds were identified by NMR spectroscopy and mass spectrometry, the absolute configuration of 3 was determined by comparison of experimental and ab initio-calculated chiroptical data. The antimicrobial activity of the bacterial extract and the isolated compounds were assayed using a set of microorganisms, and cytotoxic activities were determined against 36 human cancer cell lines.

Karamomycins A-C: 2-Naphthalen-2-yl-thiazoles from Nonomuraea endophytica.

Karamomycins A-C (2-4), the first natural 2-naphthalen-2-yl-thiazole derivatives, were isolated along with a plausible precursor molecule, 1-hydroxy-4-methoxy-2-naphthoic acid (1), uracil, 1-acetyl-ß-carboline, and actinomycin C2 from the culture broth of the terrestrial actinomycete strain GW58/450, identified as Nonomuraea endophytica. These compounds were characterized by analysis of their NMR and mass spectrometry (MS) data; the absolute configurations of 2 and 4 were determined by comparison of 13C NMR, NOESY, and circular dichroism (CD) spectra with density functional theory (DFT)-calculated data. In karamomycin C (4), the thiazole of 2 is connected to an unusual iminothiazolo[4,3- c][1,4]thiazepinone, for which we proposed a biosynthetic origin from two cysteine residues. It is closely related to ulbactin F; however, the heterocycle is enantiomeric to the latter and connected to phenol instead of 4-methoxy-1-naphthol. Karamomycins A (2) and C (4) were cytotoxic.

Studies on the constituents of Helleborus purpurascens: analysis and biological activity of the aqueous and organic extracts.

In Southeast Europe, the ethnomedicinal use of Helleborus species has a very long tradition. Cardiac steroids (Hellebrin), cysteine-rich proteins (Hellethionins) and several steroidal saponins have been identified in these plants. Aim of the present work was to investigate the amino acid composition of native extracts from the root and rootstock of Helleborus purpurascens. The amino acids have been identified by the GC-MS technique on the previously derivatised (Phenomenex Faast Kit) extract samples by comparison with the mass spectra and retention-time of the standards. A remarkable finding was a relatively intensive peak attributed to the non-proteinogenic Pipecolic acid (Pic). A cyclisation of the derivatised glutamine was observed during the GC measurement and a mechanistic pathway is described. Samples of the extract and of some isolated fractions have also been tested on; altogether 12 cancer cell lines aimed to identify further potentially cytostatic components which should be less toxic than Hellebrin. The finding of one Hellebrin-free fraction (IC50 = 0.007 mg/L) with higher cytotoxicity than Hellebrin (IC50 = 0.008 mg/L) is remarkable.

Lung Cancer: EGFR Inhibitors with Low Nanomolar Activity against a Therapy-Resistant L858R/T790M/C797S Mutant.

The treatment of non-small-cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) inhibitors is made challenging by acquired resistance caused by somatic mutations. Third-generation EGFR inhibitors have been designed to overcome resistance through covalent binding to the Cys 797 residue of the enzyme, and these inhibitors are effective against most clinically relevant EGFR mutants. However, the high dependence of these recent EGFR inhibitors on this particular interaction means that additional mutation of Cys 797 results in poor inhibitory activity, which leads to tumor relapse in initially responding patients. A new generation of irreversible and reversible mutant EGFR inhibitors was developed with strong noncovalent binding properties, and these compounds show high inhibitory activities against the cysteine-mutated L858R/T790M/C797S EGFR.

Marine Bacteria, XLVII - Psychrotolerant Bacteria from Extreme Antarctic Habitats as Producers of Rare Bis- and Trisindole Alkaloids.

From the gastrointestinal tract of a fish dredged near the South Orkney Islands in Antarctica, we isolated the psychrotolerant bacterial strain T262, which belongs to the species Vibrio splendidus. Investigation of this strain led to the isolation of a series of 15 bis- and trisindole derivatives. Among them, six new indole alkaloids, namely, turbomycin C [4'-n-butoxyphenyl-bis(1H-indol-3-yl)methane, 1a], turbomycin D [4'-n-propoxyphenyl-bis(1H-indol-3-yl)methane, 1b], turbomycin E [4'-ethoxyphenyl-bis(1H-indol-3-yl)methane, 1c], turbomycin F [4'-methoxy-3',5'-dinitrophenyl-bis(1H-indol-3-yl)methane, 2], trisindolal (3a), and 4-(1H-indol-3-yl-sulfanyl)phenol (4). Another new bisindole derivative elucidated as 2-(indol-3-ylmethyl)-indol-3-ylethanol (7a) was obtained together with six known compounds from the psychrotolerant Arthrobacter psychrochitiniphilus strain T406, isolated from the excrement of penguins. Some of the isolated compounds showed activity against both gram-positive and gram-negative bacteria at 10 µg/paper disk. Trisindolal (3a) was active against the peronosporomycetes Botrytis cinerea and Phytophthora infestans, and some of the indole derivatives indicated promising cytotoxicity towards human tumor cell lines. By exhibiting a mean IC50 of 0.45 µg/mL (1.17 µM), trisindolal (3a) showed pronounced potency and selectivity in a panel of 11 human tumor cell lines derived from 10 different tumor histotypes.

N-heterocyclic carbenes (NHC) with 1,2,4-oxadiazole-substituents related to natural products: synthesis, structure and potential antitumor activity of some corresponding gold(I) and silver(I) complexes.

This work presents the synthesis, characterization and application of eleven new gold (I) complexes 13-23 with 1,2,4-oxadiazole-containing N-heterocyclic carbene (NHC) ligands and of the NHC silver(I) complex 24. The 1,2,4-oxadiazole unit, which can be found in a variety of biologically active natural products such as phidianidines or quisqualic acid, was incorporated, along with a variety of other biologically active moieties (anthracene, indole, 2-pyridine, 2,3,4,5-tetra-O-acetyl-D-glucopyranose, quincorine and quincoridine), in order to change the lipophilicity of the complexes, so that the transport of the active units (M-NHC) though the cell wall barrier is facilitated. The biological activity of the complexes was investigated. In vitro assessment of anti-tumor activity in a panel of 12 human tumor cell lines by a monolayer assay revealed impressive potency (mean IC50 < 0.1 µM) and tumor selectivity for 6 compounds, with individual IC50 values in the low nanomolar range. The solid state structures of compounds 13, 14, 15, 17, 18, 19 and 24 were determined by X-ray diffraction analyses.

Anticancer activity of new depsipeptide compound isolated from an endophytic fungus.

A novel depsipeptide (PM181110) was purified from an endophytic fungus Phomopsis glabrae isolated from the leaves of Pongamia pinnata (family Fabaceae). The chemical structure of PM181110 was elucidated using physiochemical properties, 2D NMR and other spectroscopic methods. PM181110 is very close in structure to FE399. The compound exhibited in vitro anticancer activity against 40 human cancer cell lines with a mean IC50 value of 0.089 µM and ex vivo efficacy towards 24 human tumor xenografts (mean IC50=0.245 µM).

Total synthesis and cytotoxicity of the marine natural product malevamide D and a photoreactive analog.

The marine natural product malevamide D from the cyanobacterium Symploca hydnoides was synthesized for the first time. The final peptide coupling linked the dolaisoleuine and dolaproine subunits. The phenyl group of malevamide D was also functionalized with a photoreactive diazirine moiety, which was carried through seven reaction steps. Comprehensive assessment of the cytotoxicity in a panel of 42 human cancer cell lines revealed a geomean IC70 value of 1.5 nM (IC50 0.7 nM) for malevamide D, whereas the photoreactive derivative proved to be less active by a factor of at least 200. COMPARE analysis indicated tubulin interaction as likely mode of action of malevamide D.

Pharmacogenomics of cantharidin in tumor cells.

Cantharis vesicatoria (blister beetle) is used in Chinese medicine and has been categorized as highly toxic in the Chinese pharmacopeia. In Europe, Cantharis patches have been used since ages to treat various skin-related diseases. We investigated the cytotoxicity of the Cantharis ingredient, cantharidin, in 41 tumor cell lines (Oncotest panel) and compared the results with those of 60 cell lines of the National Cancer Institute, USA. We found profound activity at low micromolar concentrations (log 10IC50 values between -6.980 and 5.009 M). Cantharidin bound to protein phosphatase 2A (PP2A) with higher affinity (-8.12 kcal/mol) than to PP1 (-6.25 kcal/mol) in molecular docking analyses. Using a PCR array for 84 apoptosis genes, cantharidin treatment upregulated gene expression of caspase-1 and nerve growth factor receptor, but downregulated mRNA expression of Bcl-2 like protein 10, Fas ligand, and tumor necrosis factor-α. By using COMPARE analysis of microarray-based transcriptome-wide mRNA expressions, 21 genes were found to significantly correlate with response of 60 tumor cell lines to cantharidin. As shown by hierarchical cluster analysis and chi-squared test, the distribution of cell lines in the dendrogram according to their gene expression profiles predicted sensitivity or resistance to cantharidin (P=6.482 × 10(-5)). The compassionate use of Cantharis patches in two patients suffering from basalioma and Mycosis fungoides, respectively, considerably improved the diseases without signs of toxicity. In conclusion, these results indicate that cantharidin may be a useful candidate to develop novel strategies for cancer therapy.

Synthesis and characterization of novel bioactive 1,2,4-oxadiazole natural product analogs bearing the N-phenylmaleimide and N-phenylsuccinimide moieties.

Taking into consideration the biological activity of the only natural products containing a 1,2,4-oxadiazole ring in their structure (quisqualic acid and phidianidines A and B), the natural product analogs 1-(4-(3-tert-butyl-1,2,4-oxadiazol-5-yl)phenyl)pyrrolidine-2,5-dione (4) and 1-(4-(3-tert-butyl-1,2,4-oxadiazol-5-yl)phenyl)-1H-pyrrole-2,5-dione (7) were synthesized starting from 4-(3-tert-butyl-1,2,4-oxadiazol-5-yl)aniline (1) in two steps by isolating the intermediates 4-(4-(3-tert-butyl-1,2,4-oxadiazol-5-yl)phenylamino)-4-oxobutanoic acid (3) and (Z)-4-(4-(3-tert-butyl-1,2,4-oxadiazol-5-yl)phenylamino)-4-oxobut-2-enoic acid (6). The two natural product analogs 4 and 7 were then tested for antitumor activity toward a panel of 11 cell lines in vitro by using a monolayer cell-survival and proliferation assay. Compound 7 was the most potent and exhibited a mean IC50 value of approximately 9.4 µM. Aniline 1 was synthesized by two routes in one-pot reactions starting from tert-butylamidoxime and 4-aminobenzoic acid or 4-nitrobenzonitrile. The structures of compounds 1, 2, 4, 5 and 6 were confirmed by X-ray crystallography.

Doubly prenylated tryptamines: cytotoxicity, antimicrobial activity and cyclisation to the marine natural product flustramine A.

The marine natural product flustramine A was synthesised via oxidative cyclisation of Nb-methylated 1-prenyl-2-tert-prenyl-6-bromotryptamine and subsequent reduction of the resulting amidinium salt. Only the tert-prenyl group migrated, whereas the 1-prenyl group remained in place. Interestingly, the 2-tert-prenylated precursor revealed to be the biologically most active of our entire series of 21 compounds. Required for cytotoxicity and antimicrobial activity was the presence of a non-cyclised tryptamine side chain carrying a free secondary amine, whereas the presence of a 6-bromo substituent did not enhance cytotoxicity. In a panel of 42 human tumor cell lines, most sensitive were the lung and mammary cancer cell lines LXFA629L (IC50 1.9 µM) and MAXF401NL (IC50 2.4 µM), respectively. In a serial dilution assay, satisfying IC50 values of 5.9 µM against Micrococcus luteus and 7.7 µM each against Mycobacterium phlei were determined for Nb-methyl-1-prenyl-2-tert-prenyl-6-bromotryptamine.

Synthesis and antiproliferative activity of 2,5-bis(3'-indolyl)pyrroles, analogues of the marine alkaloid nortopsentin.

2,5-bis(3'-Indolyl)pyrroles, analogues of the marine alkaloid nortopsentin, were conveniently prepared through a three step procedure in good overall yields. Derivatives 1a and 1b exhibited concentration-dependent antitumor activity towards a panel of 42 human tumor cell lines with mean IC50 values of 1.54 µM and 0.67 µM, respectively. Investigating human tumor xenografts in an ex-vivo clonogenic assay revealed selective antitumor activity, whereas sensitive tumor models were scattered among various tumor histotypes.

Anticancer properties of distinct antimalarial drug classes.

We have tested five distinct classes of established and experimental antimalarial drugs for their anticancer potential, using a panel of 91 human cancer lines. Three classes of drugs: artemisinins, synthetic peroxides and DHFR (dihydrofolate reductase) inhibitors effected potent inhibition of proliferation with IC50s in the nM- low µM range, whereas a DHODH (dihydroorotate dehydrogenase) and a putative kinase inhibitor displayed no activity. Furthermore, significant synergies were identified with erlotinib, imatinib, cisplatin, dasatinib and vincristine. Cluster analysis of the antimalarials based on their differential inhibition of the various cancer lines clearly segregated the synthetic peroxides OZ277 and OZ439 from the artemisinin cluster that included artesunate, dihydroartemisinin and artemisone, and from the DHFR inhibitors pyrimethamine and P218 (a parasite DHFR inhibitor), emphasizing their shared mode of action. In order to further understand the basis of the selectivity of these compounds against different cancers, microarray-based gene expression data for 85 of the used cell lines were generated. For each compound, distinct sets of genes were identified whose expression significantly correlated with compound sensitivity. Several of the antimalarials tested in this study have well-established and excellent safety profiles with a plasma exposure, when conservatively used in malaria, that is well above the IC50s that we identified in this study. Given their unique mode of action and potential for unique synergies with established anticancer drugs, our results provide a strong basis to further explore the potential application of these compounds in cancer in pre-clinical or and clinical settings.

Novel inhibitors of epidermal growth factor receptor: (4-(Arylamino)-7H-pyrrolo[2,3-d]pyrimidin-6-yl)(1H-indol-2-yl)methanones and (1H-indol-2-yl)(4-(phenylamino)thieno[2,3-d]pyrimidin-6-yl)methanones.

Several members of the quinazoline class of known tyrosine kinase inhibitors are approved anticancer agents, often showing selectivity for receptors of the HER/ErbB-family. Combining structural elements of this class with the bisindolylmethanone-structure led to a series of novel compounds. These compounds inhibited EGFR in the nanomolar range. Moreover, inhibition of EGFR autophosphorylation in intact A431 cells was shown, with IC(50) values ranging form 0.3-1µM for compound 42, and 0.1-0.3µM for 45. In a panel of 42 human tumor cell lines the sensitivity profile of the novel compounds was shown to be similar to that of the quinazoline class of tyrosine kinase inhibitors lapatinib and erlotinib (Tarceva®).

Mechanistic studies on two dinuclear organogold(III) compounds showing appreciable antiproliferative properties and a high redox stability.

Two dinuclear oxo-bridged organogold(III) compounds, namely [(N,N,C)(2)Au(2)(µ-O)][PF(6)](2) (with N,N,CH = 6-(1-methylbenzyl)-2,2'-bipyridine, Au(2)O1; or 6-(1,1-dimethylbenzyl)-2,2'-bipyridine, Au(2)O2), were previously prepared and characterised. Their solution chemistry under physiological-like conditions has been investigated here as well as their in vitro antiproliferative properties. Notably, these compounds reveal a marked redox stability even in the presence of effective biological reductants such as ascorbic acid and glutathione. The two dinuclear gold(iii) compounds were evaluated for cytotoxic actions against a representative panel of 12 human tumor cell lines, in comparison to respective mononuclear parent compounds [(N,N,C)AuOH][PF(6)], and appreciable biological activity could be highlighted. The reactions of Au(2)O1 and Au(2)O2 with a few model proteins were studied and the ability to form metallodrug-protein adducts monitored through ESI MS methods. Typical adducts were identified where the protein is associated to monometallic gold fragments; in these adducts gold remains in the oxidation state +3 and conserves its organic ligand. A direct comparison of the biological profiles of these binuclear organogold(III) compounds with those previously reported for a series of dinuclear oxo-bridged complexes [(N,N)(2)Au(2)(µ-O)(2)][PF(6)](2) (N,N = 6(6')-substituted 2,2'-bipyridines) named Auoxo's was carried out. It emerges that the greater cytotoxicity of the latter is mainly due to the greater oxidising power of their gold(III) centres and to propensity to generate gold(i) species; in contrast, the here described bimetallic organogold(III) complexes manifest a far higher redox stability in the biological milieu coupled to lower, but still significant, antiproliferative properties. Different molecular mechanisms are thus hypothesised for these two classes of dinuclear gold(III) agents.

Molecular editing and assessment of the cytotoxic properties of iejimalide and progeny.

Systematic variation of all substructures embedded into the framework of iejimalide B (2) led to a panel of synthetic analogues of this polyunsaturated macrolide, featuring structural modifications that are not accessible by derivatization of the natural lead compound itself. The assessment of the cytotoxicity of these compounds with the aid of a monolayer proliferation assay (12 human tumor cell lines in vitro) as well as a colony formation assay (24 human tumor xenografts ex vivo) revealed the exceptional potency of 2 and several of its synthetic congeners, with IC(50) values in the picomolar range for the most sensitive cell lines. Whereas structural modifications of the macrocycle or of the side chain generally lead to a decrease in activity, changes of the peptidic terminus are not only well accommodated but engender increased tumor selectivity as well. 2 and two of the most promising analogues were selected for in vivo studies in mice, in which their activity against human tumor xenografts of breast cancer (MAXF 401) and prostate cancer (PRXF PC-3M) was tested. In contrast to a previous report in the literature however, which had claimed in vivo activity for the parent iejimalides, these tests did not show a significant therapeutic effect against the chosen solid tumors upon intraperitoneal administration.

Histone deacetylase inhibitor MGCD0103 synergizes with gemcitabine in human pancreatic cells.

Histone deacetylase inhibitors are a group of recently developed compounds that modulate cell growth and survival. We evaluated the effects of the histone deacetylase inhibitor MGCD0103 on growth of pancreatic carcinoma models following single agent treatment and in combination with gemcitabine. MGCD0103 inhibited tumor cell growth and acted synergistically with gemcitabine to enhance its cytotoxic effects. Gene expression analysis identified the cell cycle pathway as one of the most highly modulated gene groups. Our data suggest that MGCD0103 + gemcitabine might be an effective treatment for gemcitabine-refractory pancreatic cancer.

Characterization of the anticancer properties of monoglycosidic cardenolides isolated from Nerium oleander and Streptocaulon tomentosum.

AIM OF THE STUDY: For identification of the active constituents we investigated the anticancer activity of cardenolides from Streptocaulon tomentosum Wight & Arn. (Asclepiadaceae) and from Nerium oleander L. (Apocynaceae) which are both used against cancer in the traditional medicine in their region of origin. MATERIAL, METHODS AND RESULTS: The antiproliferative activity of cardenolides isolated from roots of Streptocaulon tomentosum (IC(50)<1-15.3 µM after 2 days in MCF7) and of cardenolide containing fractions from the cold aqueous extract of Nerium oleander leaves ("Breastin", mean IC(50) 0.85 µg/ml in a panel of 36 human tumor cell lines), their influence on the cellular viability and on the cell cycle (block at the G2/M-phase or at the S-phase in tumor cells, respectively) were determined using different cell lines. The murine cell line L929 and normal non-tumor cells were not affected. Bioactivity guided fractionation of Breastin resulted in the isolation of the monoglycosidic cardenolides oleandrine, oleandrigeninsarmentoside, neritaloside, odoroside H, and odoroside A (IC(50)-values between 0.010 and 0.071 µg/ml). CONCLUSIONS: The observed anticancer activities of extracts and isolated cardenolides are in agreement with the ethnomedicinal use of Streptocaulon tomentosum and Nerium oleander. The most active anticancer compounds from both species are monoglycosidic cardenolides possessing the 3ß,14ß-dihydroxy-5ß-card-20(22)-enolide structure with or without an acetoxy group at C-16. The results indicate that the cytotoxic effects are induced by the inhibition of the plasma membrane bound Na(+)/K(+)-ATPase.