The combination of yondelis and cisplatin is synergistic against human tumor xenografts.

Yondelis (trabectidin, ET-743) is a marine natural product that has shown activity both in preclinical systems and in human malignancies such as soft tissue sarcoma and ovarian cancers that are resistant to previous chemotherapies. Molecular pharmacological studies indicated that Yondelis interacts with DNA and DNA repair systems in a way that is different from Cisplatin (DDP). The current study was designed to investigate the effects of the combination of Yondelis and DDP in human cancer cell lines and in xenografts derived from different tumours. The in vitro studies performed in human TE-671 rhabdomyosarcoma, Igrov-1 and 1A9 human ovarian carcinoma cell lines showed additive effects or slight synergism. Several human tumour xenografts, such as TE-671 rhabdomyosarcoma, SK-N-DX neuroblastoma, FADU head and neck, LX-1 non-small cell lung cancer (NSCLC), H-187 melanoma and SKOV HOC 8 ovarian carcinoma, showed an antitumour effect for the combination that was greater than that of each drug when given as a single agent. No consistent changes in the activity were observed if Yondelis and DDP were given 1 h apart in sequence or simultaneously. An orthotopically transplanted human ovarian cancer HOC 8 growing in the peritoneal cavity of nude mice was used that is insensitive to Yondelis alone and only moderately sensitive to DDP alone. The combination of the two drugs produced a dramatic increase of survival lasting several months. In conclusion, the combination of Yondelis and DDP is synergistic in vivo (i.e. the antitumour effect is greater than that of each drug used as a single agent at the maximum tolerated dose (MTD)) in different human tumour xenografts. The two drugs can be combined at the MTD of each drug, thus indicating there are no overlapping toxicities. These results provide a rationale for testing the combination of Yondelis and DDP in the clinic.

No supermassive black hole in M33?

We observed the nucleus of M33, the third-brightest galaxy in the Local Group, with the Space Telescope Imaging Spectrograph at a resolution at least a factor of 10 higher than previously obtained. Rather than the steep rise expected within the radius of gravitational influence of a supermassive black hole, the random stellar velocities showed a decrease within a parsec of the center of the galaxy. The implied upper limit on the mass of the central black hole is only 3000 solar masses, about three orders of magnitude lower than the dynamically inferred mass of any other supermassive black hole. Detecting black holes of only a few thousand solar masses is observationally challenging, but it is critical to establish how supermassive black holes relate to their host galaxies, and which mechanisms influence the formation and evolution of both.

A distance to the galaxy NGC4258 from observations of Cepheid variable stars.

Cepheid variable stars pulsate in a way that is correlated with their intrinsic luminosity, making them useful as 'standard candles' for determining distances to galaxies; the potential systematic uncertainties in the resulting distances have been estimated to be only 8-10%. They have played a crucial role in establishing the extragalactic distance scale and hence the value of the Hubble constant. Here we report observations of Cepheids in the nearby galaxy NGC4258; the distance calculated from the Cepheids is 8.1 +/- 0.4 Mpc, where the uncertainty does not include possible systematic errors. There is an independently determined geometric distance to this galaxy of 7.2 +/- 0.5 Mpc, based on the observed proper motions of water masers orbiting the central black hole; the distances differ by 1.3sigma. If the maser-based distance is adopted and the Cepheid distance scale revised accordingly, the derived value of the Hubble constant would increase by 12 +/- 9%, while the expansion age of the Universe would decrease by the same amount.

Characterization of cCel3, a member of the pepper endo-beta-1,4-glucanase multigene family.

In pepper plants the enzyme endo-beta-1,4-glucanase (EGase) is encoded by a multigene family. Here is described the characterization of cCel3, a cDNA which codes for the third EGase known so far in this plant. The known members of this family are present as single copy genes, as demonstrated by a Southern analysis of the genomic DNA. Analysis of the expression of cCel3 demonstrates that the highest levels of the cCel3 mRNA are found in abscission zones of leaves and flowers activated by treatments with the plant hormone ethylene. However, the amounts of the cCel3 transcripts (detectable only by RT-PCR) are always much lower than those of cCel2 so that the latter can be regarded as the "abscission" EGase while cCel3 is likely to perform a role ancillary to that of cCel2.

Secretion, purification and activity of two recombinant pepper endo-beta-1,4-glucanases expressed in the yeast Pichia pastoris.

Two pepper endo-beta-1,4-glucanases, involved in fruit softening (cCel1) and leaf and flower abscission (cCel2), have been expressed in Pichia pastoris. Secretion was obtained by using either the mouse alpha-factor signal (cCel1) or the native signal sequence (cCel2). Times for optimal expression of the two proteins were different and cCel2 appeared very sensitive to proteolytic degradation. A one-step purification protocol yielded cCel2 in a pure form, while an additional chromatography step was necessary to purify cCel1. The two recombinant proteins are highly active and able to degrade carboxymethylcellulose in viscometric assays. Moreover, they have both a molecular mass (54 kDa) and an isoelectric point (7.2 for cCel2 and 8.5 for cCel1) equal to those of the native proteins, thus suggesting that post-translational modifications have properly occurred.

Characterization of ppEG1, a member of a multigene family which encodes endo-beta-1,4-glucanase in peach.

Three cDNA clones (pCel 10, pCel 20 and pCel 30), each encoding different endo-beta-1,4-glucanases in peach, were obtained by RT-PCR and their expression investigated by northern analysis during leaf and fruit abscission and during fruit development. This analysis allowed the detection of only the pCel 10-related mRNA. A 2.2 kb transcript accumulated in ethylene activated abscission zones of leaves and fruits, and ppEG1 (Prunus persica endoglucanase 1) the gene coding for pCel 10, was isolated and characterized. A cDNA (termed pCel 1), containing the entire open reading frame of ppEGC1, was obtained and its sequence used to define the structure of the gene and the exon/intron boundaries. ppEG1 consists of 7 exons and encodes a 497 amino acid polypeptide including a putative signal peptide at the N-terminus. The similarity of this peach endo-beta-1,4-glucanase (EGase, EC 3.2.1.4) is high (76.3%) with the ripening avocado and low (47.3%) with the bean abscission EGase. A 1639 bp region at the 5' of the transcription start site shows regulatory functions in transgenic tobacco plants, as judged by its ability to drive GUS expression in cell separation-related events.

Differential ethylene-inducible expression of cellulase in pepper plants.

Ethylene promotes the abscission of leaves and the ripening of fruits in pepper plants, and in both events an increase in cellulase activity is observed. However, two enzyme isoforms (pI 7.2 and 8.5, respectively) are differentially involved in the two physiological phenomena. The pI 8.5 form has been purified from ripe fruits. It is a glycoprotein with an apparent molecular mass of 54 kDa. Two short peptides were sequenced and a very high homology to a tomato cellulase was observed. Polyclonal antibodies, raised against the purified enzyme, have allowed us to demonstrate that the observed ethylene-induced increase in cellulase activity is paralleled by de novo synthesis of protein. Three cDNAs (CX1, CX2 and CX3), encoding different cellulases, were obtained and characterized and their expression investigated. Accumulation of all three mRNAs is induced by ethylene treatment, though to different levels. CX1 is mainly expressed in ripe fruits while CX2 is especially found in abscission zones. CX3 accumulates at very low levels in activated abscission zones. Comparisons with other known cellulases demonstrate clear heterogeneity within the higher plant cellulases. Differences in ethylene inducibility and molecular structure suggest different physiological roles for cellulase in pepper plants.