Geldanamycin, an inhibitor of the chaperone activity of HSP90, induces MAPK‐independent cell cycle arrest

Abstract

The effects of GA, an ansamycin antibiotic in development as a lead anticancer drug, were studied in mouse BP-A31 fibroblasts and in human cancer-derived cell lines. GA and related molecules act by inhibiting the chaperone function of the Hsp90 protein through competition for ATP binding. The antiproliferative effects of GA have been attributed to destabilization of the Raf-1 protein, one of the targets of Hsp90, and to the resulting inhibition of MAPK. Addition of GA to BP-A31 cells, synchronously progressing through the G 1 phase, inhibited Rb hyperphosphorylation and G 1 /S transition irrespective of the time of addition. The G 1 arrest was accompanied by a progressive decrease in Raf-1 content, especially of the phosphorylated form; however, GA caused only partial inhibition of MAPK phosphorylation. We show that GA triggers a rapid and marked decrease in the kinase activity of the cyclin E/cdk2 complex coupled with a decline in both total and cdk2-associated cyclin E. In transient transfection experiments, inhibition of cyclin E expression by GA was correlated with inhibition of the transcriptional activity of the cyclin E gene promoter. Transduction of mitogenic signals from the plasma membrane to the nucleus is relayed by the GTP-binding Ras protein and the Raf-1 protein kinase. 1,2 Activated Raf-1 phosphorylates MAPK/ ERK kinases (MEKs), 3,4 which in turn activate MAPKs/ERKs, 5 favoring their nuclear translocation 6 and consequently the activation of transcription factors. 7 Thereafter, the sequential activation of several cyclin/cdk complexes 8 controls progression through the G 1 phase toward the S phase. Cdk activity is regulated by both the cyclin abundance 9,10 and the interplay of kinases and phosphatases. [11][12][13] Cyclin D expression and assembly with cdk4/cdk6 is promoted by MAPK, thus providing a link between the 2 cascades. 14 -17 Cdk4 and cdk6 associated with D-type cyclins are activated from mid-to late G 1 phase, while cdk2 associated with cyclin E is activated in late G 1 phase. 18 Subsequently, cdk2 and cdk1 assemble with A and B cyclins and are active during the S/G 2 and M phases. Two families of specific cki proteins also modulate the activity of cyclin/cdk complexes. The INK4 family (p15, p16, p18 and p19) specifically targets the kinases associated with the D-type cyclins, whereas the Cip/Kip family (p21Kip1 , p57 Kip2 ) interacts with all cdks. 19,20 ckis of the Cip/Kip family also function as adapters, allowing the assembly and subcellular addressing of the cyclin D/cdk complexes. 17,21 Sequestration of Cip/Kip proteins by cyclin D/cdk complexes lowers the constraint of these ckis on cyclin E/cdk2 complexes. 20 The retinoblastoma gene product, Rb, and related proteins, p107 and p130, are among the key cdk substrates. Hypophosphorylated Rb is a negative regulator of the E2F family of transcription factors. 21,22 Phosphorylation of Rb occurs in an orderly manner: first, cyclin D-activated cdk4 phosphorylates Rb on specific residues (including Ser 780 ). [23][24][25] This releases t...

Akacid-medical-formulation, a novel biocidal oligoguanidine with antitumor activity reduces S-phase in prostate cancer cell lines through the Erk 1/2 mitogen-activated protein kinase pathway

“…Yim et al showed that, in DU-145 cells, treatment with high micromolar concentrations of a coumarin compound decursin yielded a decrease in cdk 2, cdk 4, cdk 6, and cyclin D1 expression and a reduction of percentage of cells in S-phase (31). Similar data were obtained in DU-145 cells with the antibiotic geldanamycin, a drug whose target is the chaperone heat shock protein 90 (32). At present, molecular mechanism of the geldanamycin effect, in particular requirement of functional tumor suppressor retinoblastoma, is not fully understood (33).…”

Activation of a novel Bcr/Abl destruction pathway by WP1130 induces apoptosis of chronic myelogenous leukemia cells

“…WP1130-induced degradation of Bcr/Abl is independent of the Hsp90/Hsp70 pathway, which engages a proteasomal-dependent mechanism for Bcr/Abl down-regulation by geldanamycin. 38,39 By targeting the Bcr/Abl protein through a distinct mechanism, WP1130 has an advantage over imatinib mesylate in that its activity is not inhibited by a variety of Abl kinase mutations, including T315I ( Figure 3A). We found that WP1130 strongly inhibited the growth of cell lines expressing either wild-type or the T315I mutant form of Bcr/Abl as well as leukemic blast progenitor cells from patients who had developed imatinib mesylate-resistant CML blast crisis ( Figure 3D).…”

“…The benzoquinone ansamycin antibiotic geldanamycin binds strongly to Hsp90 and competes for ATP binding, disrupting its chaperone activity. 38 Inactivation of Hsp90 is followed by an increase in Hsp70 expression and a shift in the binding of Bcr/Abl from Hsp90 to Hsp70, 39 which reduces the stability of Bcr/Abl. To assess the possible involvement of an Hsp90/Hsp70 mechanism in the action of WP1130, we compared the effects of WP1130 and geldanamycin on Bcr/Abl downregulation.…”

Hsp90 inhibitors cause G2/M arrest associated with the reduction of Cdc25C and Cdc2 in lung cancer cell lines

“…GA induced G1 arrest associated with decrease of both total and Cdk2-associated cyclin E, as well as partial reduction of Cdk4 in mouse fibroblast (Bedin et al 2004). In prostate cancer cell lines, 17-AAG caused the degradation of HER2, Akt, and androgen receptor, both wild and mutant forms, resulting in G1 arrest (Solit et al 2002).…”