A novel HSP90 inhibitor targeting the C-terminal domain attenuates trastuzumab resistance in HER2-positive breast cancer

Abstract

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.

HER2 Signaling and Breast Cancer Stem Cells: The Bridge behind HER2-Positive Breast Cancer Aggressiveness and Therapy Refractoriness

“…(3) inhibit tumor growth in trastuzumab-resistant JIMT-1 xenografts; and (4) reduce ALDH1 activity, the CD44 high /CD24 low subset and MFE, both in vitro and in vivo [50]. Even if no causal direct functional link between p95HER2 expression/activity and regulation of HER2+ BCSC features was clearly provided by the authors of these studies [48][49][50]74], we argue that truncated p95HER2 fragments could directly influence HER2 BC stemness.…”

“…Consistent with these important biological features, HSP90 inhibition represents an attractive strategy for the treatment of neoplasms and other diseases [ 74 ]. In this context, the C-terminal HSP90 inhibitor NCT-547 ( Table 1 ) was found to: (1) induce apoptosis and target HER2 signaling; (2) promote the degradation of FL-HER2 and truncated p95HER2 in cells properly engineered to ectopically express both molecules; (3) inhibit tumor growth in trastuzumab-resistant JIMT-1 xenografts; and (4) reduce ALDH1 activity, the CD44 high /CD24 low subset and MFE, both in vitro and in vivo [ 50 ]. Even if no causal direct functional link between p95HER2 expression/activity and regulation of HER2+ BCSC features was clearly provided by the authors of these studies [ 48 , 49 , 50 , 74 ], we argue that truncated p95HER2 fragments could directly influence HER2 BC stemness.…”

“…In particular, 611-CTF is the most potent active truncated fragment of HER2 and heavily participates in in vivo BC progression and disease relapse by triggering multiple signaling cascades that lead to a continuous activation of the PI3K/Akt pathway [ 44 ]. Dysregulation of truncated p95HER2 induced by pharmacological in vitro and in vivo targeting of HER2+ models is linked to HER2-driven stemness ( Table 1 ) [ 48 , 49 , 50 ].…”

The C-terminal HSP90 inhibitor NCT-58 kills trastuzumab-resistant breast cancer stem-like cells

“…HER2- and p95HER2-overexpressing MDA-MB-231 cells were generated using a lentiviral system, as previously described [ 51 , 52 ]. Briefly, the HER2 or p95HER2 gene was amplified by PCR [ 51 , 52 ] and then inserted into a dual promoter lentivector (CD550A-1, System Biosciences, USA). After transfection, puromycin (final concentration, 3 μg/mL) selection was performed and single colonies were isolated from a dish.…”

“…An HSP90α (C-Terminal) Inhibitor Screening Assay Kit (BPS Bioscience, CA) was used to assess recombinant C-terminal domain of HSP90α:PPID binding activity, as previously described [ 51 ]. All assays were performed with an Optiplate-384 (PerkinElmer) and measured using an AlphaScreen® microplate reader (Varioskan LUX™, Thermo Fisher Scientific, Rockford, IL).…”