Original ArticlesDCZ3112, a novel Hsp90 inhibitor, exerts potent antitumor activity against HER2-positive breast cancer through disruption of Hsp90-Cdc37 interaction
Introduction
Human epidermal growth factor receptor 2 (HER2/EGFR2/ErbB2) belongs to a family of receptor tyrosine kinases involved in regulating cell growth, differentiation, and survival [1]. HER2 is overexpressed in 25–30% of human breast cancer and is associated with high rate of recurrence and poor overall prognosis [2,3]. Thus, inhibition of the HER2 signaling pathway is a clinically established strategy for treating breast cancer. Several HER2-targeting agents, including the anti-HER2 antibodies trastuzumab [4] and pertuzumab [5,6], the EGFR/HER2 inhibitor lapatinib [7], as well as combinations of HER2-targeting agents, are currently available for treating patients with HER2-positive breast cancer [8,9]. Although these single and combinatorial regimens produce robust positive responses, the clinical benefit is usually transient, with most patients relapsing and their disease ultimately progressing after long-term treatment. Thus, there has been continuing interest in finding new HER2-targeting agents or rational drug combinations for optimal HER2 blockade to treat HER2-positive breast cancer.
Heat shock protein 90 (Hsp90) is an ATP-dependent protein chaperone involved in the regulation of cellular protein homeostasis. Hsp90 regulates the stability of client proteins, including HER2, AKT, RAF-1, CDK4 and Bcr-Abl, most of which are essential for cancer cell survival and proliferation. In fact, HER2 is one of the most well-defined client proteins in breast cancer; moreover, breast cancer is a prime target indication for Hsp90 inhibitors [[10], [11], [12], [13]]. It has been reported that Hsp90 is expressed at 2–10-fold higher levels in cancer cells than in normal cells [10]. Therefore, inhibition of HER2-mediated signaling pathways by targeting Hsp90 is a promising strategy in the treatment of HER2-positive breast cancer.
Hsp90 consists of an N-terminal domain, a middle domain and a C-terminal domain [11]. Current Hsp90 inhibitors in clinical trials all bind to the ATP pocket of the N-terminal domain and employ the same mechanism of ATP blockade to inactivate this chaperone [[12], [13], [14]]. However, none of these inhibitors has successfully reached the market owing to high toxicity and low selectivity. Thus, there is an urgent need to develop a new type of Hsp90 inhibitor distinct from the traditional strategy of inhibiting Hsp90 ATPase activity.
Hsp90 exerts its chaperone function through a process that involves transient formation of multiprotein complexes with cochaperones [15]. Disrupting interactions of Hsp90 with its cochaperones, and thereby blocking the chaperoning cycle at various stages, is also likely to achieve Hsp90 inhibition in cancer cells [16,17]. Cdc37, one of these essential cochaperones, acts through association with Hsp90 to recognize important client proteins in cancer [17]. Targeting Hsp90-Cdc37 interaction might represent a powerful strategy for treating certain kinase-dependent cancer with lower toxicity and higher selectivity compared with traditional inhibitors of Hsp90 ATPase activity. Previous studies have shown that some natural products, such as celastrol [18], withaferin A [19] and FW-04-086 [20], exhibit antitumor activity by inhibiting Hsp90 function through disruption of Hsp90-Cdc37 interaction. To date, however, no potent synthetic inhibitors of the Hsp90-Cdc37 interaction have been reported.
Our previous study showed that the designed triazine compounds targeted to N-terminal of Hsp90 exert antitumor activity by inhibiting Hsp90 function [21]. In the current study, we found that DCZ3112, a novel derivative of triazine, disrupted Hsp90-Cdc37 interaction, thereby leading to the degradation of Hsp90 client proteins. DCZ3112 showed antitumor activity predominantly in HER2-positive breast cancer and exhibited synergistic effects when combined with anti-HER2 antibodies. Importantly, DCZ3112 overcame acquired drug resistance to anti-HER2 antibodies in BT474 breast cancer cells, and also exhibited synergistic combinational effects. Thus, DCZ3112 alone or combined with anti-HER2 antibodies may constitute a targeted approach against HER2-positive breast cancer, especially for those with acquired resistance to anti-HER2 antibodies. These findings will also aid in the rational design of novel Hsp90-targeted drugs.
Section snippets
Materials
DCZ3112 was synthesized by Shanghai Institute of Materia Medica, Chinese Academy of Sciences (Shanghai, China). Geldanamycin (GM) was purchased from Sangon Biotech (Shanghai, China). Trastuzumab and pertuzumab was purchased from Roche (Basel, Switzerland).
Appropriate antibodies specific for Cdc37, HER2, AKT, CDK4, CDK6, PARP, Caspase-3, Cleaved caspase-3, Caspase-8, Caspase-9, Rb, P-Rb, Cyclin D, P-AKTS473, P-ERK1/2, and EGFR were purchased from Cell Signaling Technology (Beverly, MA, USA).
DCZ3112 binds to the N-terminal domain of Hsp90 and disrupts Hsp90-Cdc37 interaction
Screening serial triazine derivatives using SPR assays led to the identification of DCZ3112 as a novel Hsp90 inhibitor (Fig. 1A). Like the Hsp90 inhibitor GM, DCZ3112 directly bound to recombinant human full-length Hsp90α in a concentration-dependent manner, exhibiting a dissociation constant (KD) of 4.98 μM (Fig. 1B). We next employed a fluorescence polarization assay, a classic method for screening Hsp90 inhibitor binding to the ATP pocket. Unexpectedly, DCZ3112 failed to compete with FITC-GM
Discussion
HER2 is one of the most sensitive client proteins of Hsp90, and Hsp90 expression is significantly higher in breast cancer cells than in normal cells [27,28]. Accordingly, Hsp90 inhibitors have been considered potential therapeutic agents for HER2-positive breast cancer, and there is some evidence that these agents exert antitumor activity [29,30]. However, the low selectivity and high toxicity of traditional Hsp90 inhibitors and the “protective” heat shock response to them limit their clinical
Conflicts of interest
None.
Acknowledgements
We thank Professor Chrisostomos Prodromou and Eileen Hickey for their generous gifts of pRSETA-yHsp90 and pUC-Hsp90α plasmids, respectively.
This research was supported by grants from the National Natural Science Foundation of China (№ 81273546) and the Shanghai Science and Technology Committee (№ 18DZ2293200).
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2021, Acta Pharmaceutica Sinica BCitation Excerpt :Furthermore, DCZ3112 was shown to overcome resistance to either antibody. Molecular docking studies suggest the compound operates via competitive binding to the Hsp90 NTD to displace Cdc3763. In 2019, Wang and colleagues64 used molecular dynamics simulations and mutagenesis studies to discover a novel binding interaction between Glu-47 and Gln-133 on Hsp90 and Arg-167 on Cdc37.
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These authors have contributed equally to the work.