Transcription Inhibition of Heat Shock Proteins: A Strategy for Combination of 17-Allylamino-17-Demethoxygeldanamycin and Actinomycin D

Abstract

The heat shock protein (HSP) 90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) is currently in clinical trials because of its unique mechanism of action and antitumor activity. However, 17-AAG triggers the transcription and elevation of antiapoptotic HSP90, HSP70, and HSP27, which lead to chemoresistance in tumor cells. We hypothesized that inhibiting HSP90, HSP70, and HSP27 transcription may enhance 17-AAG-induced cell death in multiple myeloma cell lines. Actinomycin D (Act D), a clinically used agent and transcription inhibitor, was combined with 17-AAG. The concentrations for 17-AAG and Act D were selected based on the target actions and plasma levels during therapy. Inducible and constitutive HSP27, HSP70, and HSP90 mRNA and protein levels were measured by real-time reverse transcription-PCR and immunoblot assays. Compared with no treatment, Act D alone decreased HSP mRNA levels in MM.1S and RPMI-8226 cell lines. Combining Act D with 17-AAG did not attenuate 17-AAG-mediated increases in transcript levels of inducible HSP70; however, constitutive HSP mRNA levels were decreased. In contrast to its effect on mRNA levels, Act D was able to abrogate 17-AAG-mediated increases in all HSP protein levels. The cytotoxicity of combined Act D and 17-AAG was assessed. Treatment with Act D alone caused <40% cell death, whereas the combination of 17-AAG and Act D resulted in an increase of cell death in both multiple myeloma cell lines. In conclusion, these results indicate that 17-AAG-mediated induction of HSP70 and HSP27 expression can be attenuated by Act D and therefore can potentially improve the clinical treatment of multiple myeloma. [Cancer Res 2009;69(9):3947-54]

Clusterin Inhibition Using OGX-011 Synergistically Enhances Hsp90 Inhibitor Activity by Suppressing the Heat Shock Response in Castrate-Resistant Prostate Cancer

“…As expected (39,40), PF-04928473 or 17-AAG induces HSF-1 transcriptional activity leading to upregulation of HSP expression. Surprisingly, we found that CLU silencing abrogates, whereas CLU overexpression enhances, Hsp90 inhibitor-induced HSF-1 transcription activity, identifying a role for CLU in the regulation of HSF-1 and the heat shock response itself.…”

“…Consistent with prior reports (28, 39), here we report that Hsp90 inhibitors induce a stress response with activation of the transcription factor HSF-1 and subsequent increased levels of Hsp90 itself, Hsp70, and CLU. This heat shock response likely enhance emergence of treatment resistance, as inhibition of transcription using actinomycin D attenuates 17-AAG-mediated Hsp70 and Hsp27 expression and potentiates the effect of 17-AAG in vitro (39). In addition, inhibition of the stress response by silencing HSF-1 increases the activity of Hsp90 inhibitors (40).…”

Inhibition of heat shock protein 27 (HspB1) tumorigenic functions by peptide aptamers

“…The Hsp90 inhibitor 17-AAG is currently used in clinical trials for its antitumor activity. However, 17-AAG triggers the transcription and elevation of anti-apoptotic Hsp27, 70 and 90, which lead to chemoresistance of cancer cells (Cervantes-Gomez et al, 2009). Interestingly, targeting Hsp27 by PAs did not modulate other Hsp levels, either in cells or in vivo (data not shown), thus validating Hsp27 as a highly promising target in oncology.…”

ATP Analog Enhances the Actions of a Heat Shock Protein 90 Inhibitor in Multiple Myeloma Cells

“…Myeloma cells (MM.1S, RPMI-8226, and U266) were either untreated or treated with 0.5 M 17-AAG alone for 8 h, a combination treatment with 10 M 8-Cl-Ado for 12 h followed by 0.5 M 17-AAG for 8 h, or 10 M 8-Cl-Ado alone for 20 h. These concentrations were selected on the basis of prior reports and are clinically achievable (Gandhi et al, 2001;Cervantes-Gomez et al, 2009). Two additional combination sequences were also evaluated.…”

“…Global RNA synthesis and global protein synthesis were measured using [ 3 H]uridine or [ 3 H]leucine incorporation (37.2 or 149Ci/mmol; Moravek Biochemicals, Brea, CA), respectively, as described before (Cervantes-Gomez et al, 2009). In brief, myeloma cells were left untreated or treated according to the drug schedule.…”