Trastuzumab resistance in HER2-positive breast cancer is associated with a poorer prognosis. HSP90 is thought to play a major role in such resistance, but N-terminal inhibitors of this target have had little success. We sought to investigate the utility of NCT-547, a novel, rationally-designed C-terminal HSP90 inhibitor in the context of overcoming trastuzumab resistance. NCT-547 treatment significantly induced apoptosis without triggering the heat shock response (HSR), accompanied by caspase-3/− 7 activation in both trastuzumab-sensitive and -resistant cells. NCT-547 effectively promoted the degradation of full-length HER2 and truncated p95HER2, while also attenuating hetero-dimerization of HER2 family members. The impairment of cancer stem-like traits was observed with reductions in ALDH1 activity, the CD24low/CD44high subpopulation, and mammosphere formation in vitro and in vivo. NCT-547 was an effective inhibitor of tumor growth and angiogenesis, and no toxic outcomes were found in initial hepatic and renal analysis. Our findings suggest that NCT-547 may have applications in addressing trastuzumab resistance in HER2-positive breast cancer.
Rationale: The heat shock protein (Hsp) system plays important roles in cancer stem cell (CSC) and non-CSC populations. However, limited efficacy due to drug resistance and toxicity are obstacles to clinical use of Hsp90 inhibitors, suggesting the necessity to develop novel Hsp90 inhibitors overcoming these limitations. Methods: The underlying mechanism of resistance to Hsp90 inhibitors was investigated by colony formation assay, sphere formation assay, western blot analysis, and real-time PCR. To develop anticancer Hsp90 inhibitors that overcome the signal transducer and activator of transcription 3 (STAT3)-mediated resistance, we synthesized and screened a series of synthetic deguelin-based compounds in terms of inhibition of colony formation, migration, and viability of non-small cell lung cancer (NSCLC) cells and toxicity to normal cells. Regulation of Hsp90 by the selected compound NCT-80 [5-methoxy-N-(3-methoxy-4-(2-(pyridin-3-yl)ethoxy)phenyl)-2,2-dimethyl-2H-chromene-6-carboxamide] was investigated by immunoprecipitation, drug affinity responsive target stability assay, binding experiments using ATP-agarose beads and biotinylated drug, and docking analysis. The antitumor, antimetastatic, and anti-CSC effects of NCT-80 were examined in vitro and in vivo using various assays such as MTT, colony formation, and migration assays and flow cytometric analysis and tumor xenograft models. Results: We demonstrated a distinct mechanism in which Hsp90 inhibitors that block N-terminal ATP-binding pocket causes transcriptional upregulation of Wnt ligands through Akt- and ERK-mediated activation of STAT3, resulting in NSCLC cell survival in an autocrine or paracrine manner. In addition, NCT-80 effectively reduced viability, colony formation, migration, and CSC-like phenotypes of NSCLC cells and their sublines with acquired resistance to anticancer drugs by inducing apoptosis and inhibiting epithelial-mesenchymal transition and the growth of NSCLC patient-derived xenograft tumors without overt toxicity. With regards to mechanism, NCT-80 directly bound to the C-terminal ATP-binding pocket of Hsp90, disrupting the interaction between Hsp90 and STAT3 and degrading STAT3 protein. Moreover, NCT-80 inhibited chemotherapy- and EGFR TKI-induced programmed cell death ligand 1 expression and potentiated the antitumor effect of chemotherapy in the LLC-Luc allograft model. Conclusions: These data indicate the potential of STAT3/Wnt signaling pathway as a target to overcome resistance to Hsp90 inhibitors and NCT-80 as a novel Hsp90 inhibitor that targets both CSCs and non-CSCs in NSCLC.
Despite the development of advanced therapeutic regimens such as molecular targeted therapy and immunotherapy, the 5-year survival of patients with lung cancer is still less than 20%, suggesting the need to develop additional treatment strategies. The molecular chaperone heat shock protein 90 (Hsp90) plays important roles in the maturation of oncogenic proteins and thus has been considered as an anticancer therapeutic target. Here we show the efficacy and biological mechanism of a Hsp90 inhibitor NCT-50, a novobiocin-deguelin analog hybridizing the pharmacophores of these known Hsp90 inhibitors. NCT-50 exhibited significant inhibitory effects on the viability and colony formation of non-small cell lung cancer (NSCLC) cells and those carrying resistance to chemotherapy. In contrast, NCT-50 showed minimal effects on the viability of normal cells. NCT-50 induced apoptosis in NSCLC cells, inhibited the expression and activity of several Hsp90 clients including hypoxia-inducible factor (HIF)-1α, and suppressed pro-angiogenic effects of NSCLC cells. Further biochemical and in silico studies revealed that NCT-50 downregulated Hsp90 function by interacting with the C-terminal ATP-binding pocket of Hsp90, leading to decrease in the interaction with Hsp90 client proteins. These results suggest the potential of NCT-50 as an anticancer Hsp90 inhibitor.
N-terminal HSP90 inhibitors in development have had issues arising from heat shock response (HSR) induction and off-target effects. We sought to investigate the capacity of NCT-58, a rationally-synthesized C-terminal HSP90 inhibitor, to kill trastuzumab-resistant HER2-positive breast cancer stem-like cells. NCT-58 does not induce the HSR due to its targeting of the C-terminal region and elicits anti-tumor activity via the simultaneous downregulation of HER family members as well as inhibition of Akt phosphorylation. NCT-58 kills the rapidly proliferating bulk tumor cells as well as the breast cancer stem-like population, coinciding with significant reductions in stem/progenitor markers and pluripotent transcription factors. NCT-58 treatment suppressed growth and angiogenesis in a trastuzumab-resistant xenograft model, concomitant with downregulation of ICD-HER2 and HSF-1/HSP70/HSP90. These findings warrant further investigation of NCT-58 to address trastuzumab resistance in heterogeneous HER2-positive cancers.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.