fun stuff (given this sharp decline at a point where the companies are committed to phase II and where 17-AAG is being investigated as a complement to MLN341, it's probably worth the effort to step up stock-related research)......
Carcinogenesis 2003 May;24(5):851-9
Inhibition of telomerase activity by geldanamycin and 17-allylamino, 17-demethoxygeldanamycin in human melanoma cells.
Villa R, Folini M, Porta CD, Valentini A, Pennati M, Daidone MG, Zaffaroni N.
Department of Experimental Oncology, Unit 10, Istituto Nazionale per lo Studio e la Cura dei Tumori, Via Venezian 1, 20133 Milan, Italy.
As it has been demonstrated that the heat shock protein 90 (HSP90) is required for the assembly and activation of telomerase in human cells, we investigated the effect exerted by the ansamycin antibiotics geldanamycin (GA) and 17-allylamino,17-demethoxygeldanamycin (17-AAG), two well-known inhibitors of the HSP90 chaperone function, on telomerase activity in JR8 human melanoma cells. Using an antibody to HSP90, we precipitated the telomerase activity associated with the molecular chaperone. The results of TRAP (telomeric repeat amplification protocol) experiments carried out on HSP90 immunoprecipitates showed that exposure to 100 ng/ml GA and 17-AAG induced a significant (P < 0.01) inhibition of telomerase activity, which was observed at earlier time points than drug-induced inhibition of cell proliferation. Superimposable results were obtained from TRAP experiments carried out on total JR8 protein extracts. To investigate whether the basal level of telomerase activity of the tumour cell system plays a role in determining the cellular response to 17-AAG, we compared the cytotoxic activity of the drug in JR8 cells and in two JR8-derived clones that were stably transfected with a hammerhead ribozyme targeting the RNA template of telomerase and were characterized by a markedly lower telomerase activity than the parental cells. The cytotoxicity results indicated that both ribozyme-transfectant clones were almost 2-fold more sensitive to 72 h 17-AAG exposure than JR8 cells as a consequence of a more than double apoptotic response [in terms of the percentage of apoptotic nuclei in cells stained with propidium iodide and the percentage of Tdt-mediated dUTP nick-end labelling (TUNEL)-positive cells]. In summary, our results suggest that (i) telomerase is a target of GA and 17-AAG action and its inhibition may contribute to the cytotoxic activity of the drugs, (ii) the basal level of telomerase activity of the tumour cell system may also have a role in influencing 17-AAG cytotoxicity.
Chem Biol 2003 Apr;10(4):361-8
Crystal Structure and Molecular Modeling of 17-DMAG in Complex with Human Hsp90.
Jez JM, Chen JC, Rastelli G, Stroud RM, Santi DV.
Kosan Biosciences, Inc., 3832 Bay Center Place, 94545, Hayward, CA, USA
Hsp90 is an attractive chemotherapeutic target because it chaperones the folding of proteins found in multiple signal transduction pathways. We describe the 1.75 A resolution crystal structure of human Hsp90alpha (residues 9-236) complexed with 17-desmethoxy-17-N,N-dimethylaminoethylamino-geldanamycin (17-DMAG). The structure revealed an altered set of interactions between the 17-substituent and the protein compared to geldanamycin and the 17-dimethylaminoethyl moiety pointing into solvent, but otherwise was similar to that reported for the complex with geldanamycin. Targeted molecular dynamics simulations and energetic analysis indicate that geldanamycin undergoes two major conformational changes when it binds Hsp90, with the key step of the conversion being the trans to cis conformational change of the macrocycle amide bond. We speculate that 17-DMAG analogs constrained to a cis-amide in the ground state could provide a significant increase in affinity for Hsp90. |
