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Cancer emergence is associated with cellular adaptations to altered signal transduction mechanisms arbitrated by mutated kinases. Since conventional kinase inhibitors can exhibit certain limitations to such kinase adaptations, overcoming kinase adaptation for cancer treatment gains importance. The cancer chaperone, Hsp90, is implicated in the conformational maturation and functional stabilization of mutated gene products. However, its role in kinase adaptations is not explored in detail. Therefore, the present study aims to understand the mechanisms of Hsp90-dependent kinase adaptation and develop a novel antitumor strategy. We chose malignant human lung cancer cells to demonstrate Hsp90-dependent RAF oncogene adaptation. We show that RAF oncogene adaptations were predominant over wild type RAF and are facilitated by conformation-specific Hsp90. Consequently, the conformation-specific Hsp90 inhibitor, 17AAG, interfered with oncogenic RAF stability and function and inhibited cell proliferation. The enforced cytostasis further triggered premature cellular senescence and acted as an efficient and irreversible tumor suppressor mechanism. Our results also display that oncogenic RAF interactions with Hsp90 require the middle-charged region of the chaperone. Our mice xenografts revealed that 17AAG pretreated tumor cells lost their ability to proliferate and metastasize in vivo. In summary, we demonstrated Hsp90-dependent kinase adaptation in tumor cells and the effect of Hsp90 inhibition in triggering premature senescence to interfere with the tumor progression. Our findings are of both biological relevance and clinical importance.

Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the main pathogens that seriously affect the sustainable development of sericulture industry. Inhibition of Hsp90 by Hsp90 inhibitor, geldanamycin (GA) significantly suppresses BmNPV proliferation in Bombyx mori, while the functional mechanism is not clear. LncRNA has been widely reported to play an important role in immune responses and host-virus interactions in mammalian. However, related research has been rarely reported on silkworm. In this study, firstly, we confirmed the decrease of BmNPV ORF75 protein in the BmNPV-infected BmN cells treated with GA. Next, by using a genome-wide transcriptome analysis, we compared the lncRNA and mRNA expression profiles in BmNPV infected BmN cells treated with or without GA and identified a total of 282 differentially expressed lncRNAs (DElncRNAs) and 523 DEmRNAs. KEGG pathway analysis revealed DEmRNA were mainly involved in ubiquitin mediated proteolysis, spliceosome, RNA transport and oxidative phosphorylation. Further, we selected 27 immune-related DEmRNAs, which displayed the similar changes of expression patterns on both protein level and transcript level to construct DElncRNA-DEmRNA network. In addition, based on the DElncRNA-bmo-miR-278-3p-BmHSC70 regulatory network, we explored the potential function of several lncRNAs as sponges to inhibit the regulatory effect of bmo-278-3p on Bombyx mori heat shock protein cognate 70 (BmHSC70). Our finding suggests that lncRNAs play a role in the regulation of BmNPV proliferation by Hsp90.

Compound 7t, 4-(4-bromophenyl)-6-(1-(4-chlorophenyl)-3-(4-nitrophenyl)-1H-pyrazol-4-yl) pyrimidin-2-amine, is a proven potent anticancer agent exhibiting Hsp90 inhibition in our previous studies. Herein, we explored the apoptotic potential of compound 7t by Annexin V assay. The mechanism underlining the apoptosis process is elucidated. As a potent Hsp90 inhibitor, compound 7t would induce the mitochondrial stress leading to increased permeability of its membrane, that would subsequently initiate the apoptosis in MCF-7 cells. This was proven by increased J-monomer formation using JC-1 stain. Moreover, due to the impaired mitochondrial function, compound 7t also exaggerated the apoptosis process by ROS generation as proved by DCFDA staining. The morphological and nuclear changes in MCF-7 cells following apoptosis were identified by AO/EB and DAPI staining techniques. It also induced subG1 phase cell cycle arrest. Thus, compound 7t could serve as potential drug in the treatment regimen of breast cancer.

Hsp90 is a ubiquitous chaperone with important roles in the organization and maturation of client proteins that are involved in the progression and survival of cancer cells. Multiple oncogenic pathways can be affected by inhibition of Hsp90 function through degradation of its client proteins. That makes Hsp90 a therapeutic target for cancer treatment. 17-allylamino-17-demethoxy-geldanamycin (17-AAG) is a potent Hsp90 inhibitor that binds to Hsp90 and inhibits its chaperoning function, which results in the degradation of Hsp90's client proteins. There have been several preclinical studies of 17-AAG as a single agent or in combination with other anticancer agents for a wide range of human cancers. Data from various phases of clinical trials show that 17-AAG can be given safely at biologically active dosages with mild toxicity. Even though 17-AAG has suitable pharmacological potency, its low water solubility and high hepatotoxicity could significantly restrict its clinical use. Nanomaterials-based drug delivery carriers may overcome these drawbacks. In this paper, we review preclinical and clinical research on 17-AAG as a single agent and in combination with other anticancer agents. In addition, we highlight the potential of using nanocarriers and nanocombination therapy to improve therapeutic effects of 17-AAG.

177Lu-octreotate is an FDA-approved radionuclide therapy for patients with gastroenteropancreatic neuroendocrine tumours (NETs) expressing somatostatin receptors. The 177Lu-octreotate therapy has shown promising results in clinical trials by prolonging progression-free survival, but complete responses are still uncommon. The aim of this study was to improve the 177Lu-octreotate therapy by means of combination therapy. To identify radiosensitising inhibitors, two cell lines, GOT1 and P-STS, derived from small intestinal neuroendocrine tumours (SINETs), were screened with 1224 inhibitors alone or in combination with external radiation. The screening revealed that inhibitors of Hsp90 can potentiate the tumour cell-killing effect of radiation in a synergistic fashion (GOT1; false discovery rate

Melanoma remains incurable skin cancer, and targeting heat shock protein 90 (HSP90) is a promising therapeutic approach. In this study, we investigate the effect of 17-aminogeldanamycin, a potent HSP90 inhibitor, on nuclear factor-kappa B (NF-&kappa, B) activity in BRAFV600E and NRASQ61R patient-derived melanoma cell lines. We performed time-lapse microscopy and flow cytometry to monitor changes in cell confluence and viability. The NF-&kappa, B activity was determined by immunodetection of phospho-p65 and assessment of expression of NF-&kappa, B-dependent genes by quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, and enzyme-linked immunosorbent assay (ELISA). Constitutive activity of p65/NF-&kappa, B was evident in all melanoma cell lines. Differences in its level might be associated with genetic alterations in CHUK, IL1B, MAP3K14, NFKBIE, RIPK1, and TLR4, while differences in transcript levels of NF-&kappa, B-inducible genes revealed by PCR array might result from the contribution of other regulatory mechanisms. 17-Aminogeldanamycin markedly diminished the level of phospho-p65, but the total p65 protein level was unaltered, indicating that 17-aminogeldanamycin inhibited activation of p65/NF-&kappa, B. This conclusion was supported by significantly reduced expression of selected NF-&kappa, B-dependent genes: cyclin D1 (CCND1), C-X-C motif chemokine ligand 8 (CXCL8), and vascular endothelial growth factor (VEGF), as shown at transcript and protein levels, as well as secretion of IL-8 and VEGF. Our study indicates that 17-aminogeldanamycin can be used for efficient inhibition of NF-&kappa, B activity and the simultaneous diminution of IL-8 and VEGF levels in the extracellular milieu of melanoma.

In genetic diseases, the most prevalent mechanism of pathogenicity is an altered expression of dosage-sensitive genes. Drugs that restore physiological levels of these genes should be effective in treating the associated conditions. We developed a screening strategy, based on a bicistronic dual-reporter vector, for identifying compounds that modulate proteins levels, and used it in a pharmacological screening approach. To provide a proof-of-principle, we chose Autosomal Dominant LeukoDystrophy (ADLD), an ultra-rare adult-onset neurodegenerative disorder caused by lamin B1 (LMNB1) overexpression. We used a stable CHO cell line simultaneously expresses an AcGFP reporter fused to LMNB1 and a Ds-Red normalizer. Using high-content imaging analysis, we screened a library of 717 biologically active compounds and approved drugs, and identified alvespimycin, an HSP90 inhibitor, as a positive hit. We confirmed that alvespimycin can reduce LMNB1 levels by 30-80% in five different cell lines (fibroblasts, NIH3T3, CHO, COS-7 and rat primary glial cells). In ADLD fibroblasts, alvespimycin reduced cytoplasmic LMNB1 by about 50%. We propose this approach for effectively identifying potential drugs for treating genetic diseases associated with deletions/duplications, and paving the way towards phase II clinical trials. This article is protected by copyright. All rights reserved.

Aim/Purpose: Exposure to high levels of hydrochloric acid (HCl) is associated with severe lung injury including both acute inflammation and chronic lung disease, which leads to the development of p...

Photodynamic therapy (PDT) ablates malignancies by applying focused near-infrared (nIR) light onto a lesion of interest after systemic administration of a photosensitizer (PS); however, the accumulation of existing PS is not tumor-exclusive. We developed a tumor-localizing strategy for PDT, exploiting the high expression of heat shock protein 90 (Hsp90) in cancer cells to retain high concentrations of PS by tethering a small molecule Hsp90 inhibitor to a PS (verteporfin, VP) to create an Hsp90-targeted PS (HS201). HS201 accumulates to a greater extent than VP in breast cancer cells both in vitro and in vivo, resulting in increased treatment efficacy of HS201-PDT in various human breast cancer xenografts regardless of molecular and clinical subtypes. The therapeutic index achieved with Hsp90-targeted PDT would permit treatment not only of localized tumors, but also more diffusely infiltrating processes such as inflammatory breast cancer., Heat shock protein 90 (Hsp90) is highly expressed in cancer cells. Kaneko et al show that targeting Hsp90 on the cancer cell surface with a photosensitizer tethered to an Hsp90 small molecule inhibitor improves the efficacy of photodynamic therapy in various human breast cancer xenografts, suggesting a therapeutic avenue for human disease.

Pyroptosis is a recently discovered inflammatory form of programmed cell death which is mostly triggered by infection with intracellular pathogens and critically contributes to inflammation. Mitigating pyroptosis may be a potential therapeutic target in inflammatory diseases. However, small chemicals to reduce pyroptosis is still elusive. In the present study, we screened 155 chemicals from a microbial natural product library and found Geldanamycin, an HSP90 inhibitor, profoundly rescued THP-1 cells from pyroptosis induced by LPS plus Nigericin treatment. Consistently, other HSP90 inhibitors, including Radicicol, 17-DMAG and 17-AAG, all ameliorated pyroptosis in THP-1 cells by suppressing the inflammasome/Caspase-1/GSDMD signal pathway in pyroptosis. HSP90 inhibition compromised the protein stability of NLRP3, a critical component of the inflammasome. Moreover, up-regulated HSP70 may also contribute to this effect. HSP90 inhibition may thus be a potential therapeutic strategy in the treatment of inflammatory diseases in which pyroptosis plays a role.

Mutation and translocation of fibroblast growth factor receptors often lead to aberrant signaling and cancer. This work focuses on the t(8;22)(p11;q11) chromosomal translocation which creates the breakpoint cluster region (BCR) fibroblast growth factor receptor1 (FGFR1) (BCR-FGFR1) fusion protein. This fusion occurs in stem cell leukemia/lymphoma, which can progress to atypical chronic myeloid leukemia, acute myeloid leukemia, or B-cell lymphoma. This work focuses on the biochemical characterization of BCR-FGFR1 and identification of novel therapeutic targets. The tyrosine kinase activity of FGFR1 is required for biological activity as shown using transformation assays, interleukin-3 independent cell proliferation, and liquid chromatography/mass spectroscopy analyses. Furthermore, BCR contributes a coiled-coil oligomerization domain, also essential for oncogenic transformation by BCR-FGFR1. The importance of salt bridge formation within the coiled-coil domain is demonstrated, as disruption of three salt bridges abrogates cellular transforming ability. Lastly, BCR-FGFR1 acts as a client of the chaperonin heat shock protein 90 (Hsp90), suggesting that BCR-FGFR1 relies on Hsp90 complex to evade proteasomal degradation. Transformed cells expressing BCR-FGFR1 are sensitive to the Hsp90 inhibitor Ganetespib, and also respond to combined treatment with Ganetespib plus the FGFR inhibitor BGJ398. Collectively, these data suggest novel therapeutic approaches for future stem cell leukemia/lymphoma treatment: inhibition of BCR oligomerization by disruption of required salt bridges; and inhibition of the chaperonin Hsp90 complex.

Although microglial reaction to heat shock is considered to be protective, heat shock is still a potential hazard caused by high temperatures. Recent studies indicate that the inhibition of the 90-kDa heat shock protein (HSP90) increasing the protective heat shock response and suppressing inflammatory signalling pathways in several diseases. Nevertheless, the effects of heat shock on microglial pro-inflammatory responses are not completely identical. Here, we aim to investigate the effect of the HSP90 inhibitor ganetespib on microglial pro-inflammatory responses following heat shock. HSP90 isoforms were determined by transfecting N9 microglial cells (N9 cells) with enzymatically prepared siRNA (esiRNAs). We found that heat shock significantly increased the secretion of tumour necrosis factor alpha (TNF-α), interleukin (IL)-1β, IL-6 and nitric oxide (NO), and the phosphorylation of extracellular signal-regulated kinase (ERK), Janus-activated kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor alpha (IκB-α) and p65 nuclear factor kappa-light-chain-enhancer of activated B cells (p65 NF-κB) in N9 cells. These increases, except for phospho-p65, were attenuated efficiently in a dose-dependent manner by ganetespib pretreatment. Furthermore, the suppression of heat shock-evoked cytokines and NO production, and the phosphorylation of ERK, JAK2 and STAT3 in cytosols and/or nuclei were also observed by administering esiRNA HSP90β, but not HSP90α, in heat shock-treated N9 cells. Taken together, our findings demonstrate that the HSP90 inhibitor ganetespib blocks pro-inflammatory responses in heat shock-treated N9 cells via a signalling mechanism involving HSP90β and STAT3.

Heat shock protein 90 (HSP90), a highly and unique chaperone, presents as a double-edged sword. It plays an essential role in many physiological and pathological processes, including tumor development. The current review highlights a recent understanding of the roles of HSP90 in molecular mechanisms underlying cancer survival and progression. HSP90 and its client proteins through the regulation of oncoproteins including signaling proteins, receptors, and transcriptional factors involved in tumorigenesis. It also has potential clinical application as diagnostic and prognostic biomarkers for assessing cancer progression. In this way, using HSP90 to develop new anticancer therapeutic agents including HSP90 inhibitors, anti-HSP90 antibody, and HSP90-based vaccines has been promising.

Anaplastic thyroid carcinoma (ATC) is a highly aggressive tumor with a poor prognosis and a low median survival rate because of insufficient effective therapeutic modalities. Recently, mesoporous silica nanoparticles (MSNs) as a green non-toxic and safe nanomaterial have shown advantages to be a drug carrier and to modify the targeting group to the targeted therapy. To aim of the study was to explore the effects of MSNs co-loading with 17-allylamino-17-demethoxy-geldanamycin (17-AAG; HSP90 inhibitor) and 9-(6-aminopyridin-3-yl)-1-(3-(trifluoromethyl)phenyl)benzo[h][1,6]naphthyridin-2(1H)-one (Torin2; mTOR inhibitor) by targeting vascular endothelial growth factor receptor 2 (VEGFR2) on the viability of human anaplastic thyroid carcinoma FRO cells. The cytotoxicity of 17-AAG and Torin2 were analyzed by MTT assay. The possible synergistic antitumor effects between 17-AAG and Torin2 were evaluated by CompuSyn software. Flow cytometry was performed to assess the VEGFR2 targeting of (17-AAG+Torin2)@MSNs-anti-VEGFR2 ab and uptake by FRO cells. An ATC xenograft mouse model was established to assess the antitumor effect of (17-AAG+Torin2)@MSNs-anti-VEGFR2 ab in vivo. The results revealed that the combination of 17-AAG and Torin2 inhibited the growth of FRO cells more effectively compared with single use of these agents. Additionally, the synergistic antitumor effect appeared when concentration ratio of the two drugs was 1:1 along with total drug concentration greater than 0.52 µM. Furthermore, in an ATC animal model, it was revealed that the (17-AAG+Torin2)@MSNs-anti-VEGFR2 ab therapy modality could most effectively prolong the median survival time [39.5 days vs. 33.0 days (non-targeted) or 27.5 days (control)]. Compared to (17-AAG+Torin2)@MSNs, the (17-AAG+Torin2)@MSNs-anti-VEGFR2 ab could not only inhibit ATC cell growth but also prolong the median survival time of tumor-bearing mice in vivo and vitro more effectively, which may provide a new promising therapy for ATC.

A series of isosteric surrogates of the 4-phenyl group in luminespib were investigated as new scaffolds of the Hsp90 inhibitor for the discovery of novel antitumor agents. Among the synthesized surrogates of isoxazole and pyrazole, compounds 4a, 5e and 12b exhibited potent Hsp90 inhibition in ATPase activity and Her2 degradation assays and significant antitumor activity in A2780 and HCT116 cell lines. Animal studies indicated that compared to luminespib, their activities were superior in A2780 or NCI-H1975 tumor xenograft models. A molecular modeling study demonstrated that compound 4a could fit nicely into the N-terminal ATP binding pocket.

About 15% of intrahepatic cholangiocarcinomas (ICCs) express constitutively active fibroblast growth factor receptor 2 (FGFR2) fusion proteins (FFs) generated by chromosomal translocations. FFs have been nominated as oncogenic drivers because administration of FGFR tyrosine kinase inhibitors (F-TKIs) can elicit meaningful objective clinical responses in patients carrying FF-positive ICC. Thus, optimization of FF targeting is a pressing clinical need. Herein, we report that three different FFs, previously isolated from ICC samples, are heat shock protein 90 (HSP90) clients and undergo rapid degradation upon HSP90 pharmacological blockade by the clinically advanced HSP90 inhibitor ganetespib. Combining catalytic suppression by the F-TKI BGJ398 with HSP90 blockade by ganetespib suppressed FGFR2-TACC3 (transforming acidic coiled-coil containing protein 3) signaling in cultured cells more effectively than either BGJ398 or ganetespib in isolation. The BGJ398 + ganetespib combo was also superior to single agents when tested in mice carrying subcutaneous tumors generated by transplantation of FGFR2-TACC3 NIH3T3 transformants. Of note, FF mutants known to enforce clinical resistance to BGJ398 in ICC patients retained full sensitivity to ganetespib in cultured cells. Conclusion: Our data provide a proof of principle that upfront treatment with the BGJ398 + ganetespib combo improves therapeutic targeting of FGFR2 fusions in an experimental setting, which may be relevant to precision medicine approaches to FF-driven ICC.

Heat-shock protein 90 (HSP90) is a molecular chaperone that activates oncogenic transformation in several solid tumors, including lung and breast cancers. Ganetespib, a most promising candidate among several HSP90 inhibitors under clinical trials, has entered Phase III clinical trials for cancer therapy. Despite numerous evidences validating HSP90 as a target of anticancer, there are few studies on PET agents targeting oncogenic HSP90. In this study, we synthesized and biologically evaluated a novel 18F-labeled 5-resorcinolic triazolone derivative (1, [18F]PTP-Ganetespib) based on ganetespib. [18F]PTP-Ganetespib was labeled by click chemistry of Ganetespib-PEG-Alkyne (10) and [18F]PEG-N3 (11) with 37.3 ± 5.11% of radiochemical yield and 99.7 ± 0.09% of radiochemical purity. [18F]PTP-Ganetespib showed proper LogP (0.96 ± 0.06) and good stability in human serum over 97% for 2 h. [18F]PTP-Ganetespib showed high uptakes in breast cancer cells containing triple negative breast cancer (TNBC) MDA-MB-231 and Her2-negative MCF-7 cells, which are target breast cancer cell lines of HSP90 inhibitor, ganetespib, as an anticancer. Blocking of HSP90 by the pretreatment of ganetespib exhibited significantly decreased accumulation of [18F]PTP-Ganetespib in MDA-MB-231 and MCF-7 cells, indicating the specific binding of [18F]PTP-Ganetespib to MDA-MB-231 and MCF-7 cells with high HSP90 expression. In the biodistribution and microPET imaging studies, the initial uptake into tumor was weaker than in other thoracic and abdominal organs, but [18F]PTP-Ganetespib was retained relatively longer in the tumor than other organs. The uptake of [18F]PTP-Ganetespib in tumors was not sufficient for further development as a tumor-specific PET imaging agent by itself, but this preliminary PET imaging study of [18F]PTP-Ganetespib can be basis for developing new PET imaging agents based on HSP90 inhibitor, ganetespib.

Human rhinoviruses (HRV) are one of the major causes of common cold in humans and are also associated with acute asthma and bronchial illness. Heat-shock protein 90 (Hsp90), a molecular chaperone, is an important host factor for the replication of single-strand RNA viruses. In the current study, we examined the effect of the Hsp90 inhibitor pochonin D, in vitro and in vivo, using a murine model of human rhinovirus type 1B (HRV1B) infection. Our data suggested that Hsp90 inhibition significantly reduced the inflammatory cytokine production and lung damage caused by HRV1B infection. The viral titer was significantly lowered in HRV1B-infected lungs and in Hela cells upon treatment with pochonin D. Infiltration of innate immune cells including granulocytes and monocytes was also reduced in the bronchoalveolar lavage (BAL) by pochonin D treatment after HRV1B infection. Histological analysis of the lung and respiratory tract showed that pochonin D protected the mice from HRV1B infection. Collectively, our results suggest that the Hsp90 inhibitor, pochonin D, could be an attractive antiviral therapeutic for treating HRV infection.

Introduction Heat shock protein 70 (Hsp70) and heat shock protein 90 (Hsp90) chaperones are indispensable to lung cancer cells for their survival and proliferation. In this study we evaluated and compared anticancer potential of methylene blue (MB) as an Hsp70 inhibitor, novobiocin (NB) a well-known Hsp90 inhibitor and their combination. Methods In vitro evaluation was done by cell viability assays, fluorescent staining, and flow cytometry analysis using A549 non-small cell lung cancer cells. In vivo anticancer activity was investigated by evaluating oxidative stress, tumor biomarkers, weight, lung microarchitecture, and Hsp70 and Hsp90 inhibitions via immunoblotting in benzo[a]pyrene induced lung carcinogenesis mice model. Results Using A549 NSCLC cells, we found MB demonstrated lower cell viability versus NB. Together, MB + NB resulted in further decrease in cell viability. SRB assay revealed significantly superior and similar potency for MB versus NB and MB + NB (1:1) versus MB, respectively. Fluorescent staining and flow cytometry analysis displayed early apoptosis by MB (11.4%); early and late apoptosis by MB + NB (13.8%). In vivo, MB significantly inhibited Hsp70. Furthermore, MB significantly alleviated tumor biomarkers (ADA and LDH) and improved lung histopathological features more than NB. Additionally, MB significantly improved SOD, not more than MB + NB or NB and improved LPO. Conclusion MB demonstrated potent anticancer activity in vitro and in vivo via inhibition of Hsp70 in benzo[a]pyrene induced lung carcinogenesis in mice.

The development of new treatments for castrate resistant prostate cancer (CRPC) must address such challenges as intrinsic tumor heterogeneity and phenotypic plasticity. Combined PTEN/TP53 alterations represent a major genotype of CRPC (25–30%) and are associated with poor outcomes. Using tumor-derived, castration-resistant Pten/Tp53 null luminal prostate cells for comprehensive, high-throughput, mechanism-based screening, we identified several vulnerabilities among >1900 compounds, including inhibitors of: PI3K/AKT/mTOR, the proteasome, the cell cycle, heat shock proteins, DNA repair, NFκB, MAPK, and epigenetic modifiers. HSP90 inhibitors were one of the most active compound classes in the screen and have clinical potential for use in drug combinations to enhance efficacy and delay the development of resistance. To inform future design of rational drug combinations, we tested ganetespib, a potent second-generation HSP90 inhibitor, as a single agent in multiple CRPC genotypes and phenotypes. Ganetespib decreased growth of endogenous Pten/Tp53 null tumors, confirming therapeutic activity in situ. Fifteen human CRPC LuCaP PDX-derived organoid models were assayed for responses to 110 drugs, and HSP90 inhibitors (ganetespib and onalespib) were among the select group of drugs (75% of models) at high potency (IC50

Internal tandem duplication (ITD) in Fms-like tyrosine kinase 3 (FLT3) is frequently observed in acute myeloid leukemia (AML). Quizartinib, gilteritinib, and midostaurin are inhibitors against FLT3-ITD that have good efficacy for FLT3-ITD-positive AML patients. Long-term administration leads to drug resistance through acquired tyrosine kinase domain (TKD) mutations in FLT3-ITD, such as N676K, F691L, D835V, and Y842C. Here, our screen to detect inhibitors capable of overcoming resistance to FLT3 inhibitors identified heat shock protein (HSP) 90 inhibitors as potential candidates. Although Ba/F3 cells expressing FLT3-ITD with TKD mutations (Ba/F3-ITD+N676K, Ba/F3-ITD+F691L, Ba/F3-ITD+D835V, and Ba/F3-ITD+Y842C) showed various resistance patterns to FLT3 inhibitors compared with Ba/F3-ITD cells that express FLT3-ITD lacking TKD mutations, they were more sensitive to HSP90 inhibitors than Ba/F3 cells. Notably, the Ba/F3-ITD+D835V cells were the most sensitive to HSP90 inhibitors. Treatment with HSP90 inhibitors downregulated FLT3 and its downstream signaling and induced G1 arrest followed by apoptosis in Ba/F3-ITD+N676K, Ba/F3-ITD+F691L, Ba/F3-ITD+Y842C, and especially Ba/F3-ITD+D835V cells at lower concentrations compared with Ba/F3-ITD cells. The downregulation of FLT3-ITD+D835V was caused by rapid proteolysis in autophagy. Similar results were also observed in the quizartinib-resistant MV4-11 cells, QR1 and QR2, which were established by culturing cells in the presence of quizartinib and harbored FLT3-ITD+D835H and FLT3-ITD+D835V, respectively, in a single allele. Interestingly, the efficacies of HSP90 inhibitors in QR cells are reversely correlated with that of quizartib, but not to gilteritinib and midostaurin. Collectively, HSP90 inhibitors are good candidates to overcome drug resistance in AML with various FLT3-ITD TKD mutations.

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.

Hepatocellular carcinoma (HCC) is a typical hypervascular solid tumor. Vasculogenic mimicry (VM) formed by aggressive tumor cells to mimic vasculogenic networks plays an important role in the tumor malignancy of HCC. Hsp90β promotes endothelial cell-dependent angiogenesis in HCC. However, the relationship between Hsp90β and VM formation is unclear. In this study, we found that Hsp90β is positively correlated with VM and EMT marker proteins in HCC tissues and promotes tube formation, cell migration, and invasion in vitro. Hsp90β interacts with Twist1 and promotes its deubiquitination and stabilization to nuclear translocation and enhances the VE-cadherin promoter activity. Results of in vitro analysis indicate that Hsp90β enhances the tumor VM in tumor-burdened mice, and the Hsp90 inhibitor NVP-BEP800 suppresses VM formation by releasing Hsp90β and Twist1 interaction. This study provides a potential antitumor therapy for inhibiting VM by targeting Hsp90β in HCC.

Acetylation plays an important role in regulating the chaperone activity of heat shock protein 90 (Hsp90) during malignant transformation through the stabilization and conformational maturation of oncogenic proteins. However, the functional acetylation sites, potential anticancer drug targets, are still emerging. We found that acetylation at K292 in Hsp90α is critical for the development and treatment of breast cancer. Acetylation at K292 not only augments the affinity of Hsp90 to ATP, cochaperones, and client proteins but it also promotes cancer cell colony formation, migration, and invasion in vitro as well as tumor growth in vivo. Importantly, K292-acetylated Hsp90 has been validated as an exciting anticancer drug target by interfering with the complex formation between K292-acetylated Hsp90 and cochaperone Cdc37, leading to diminishment of kinase client maturation and proteasome-dependent degradation of kinase substrates. Furthermore, we showed that simvastatin prevented, whereas LBH589 promoted, the progression of Hsp90 chaperone cycling and client maturation, resulting in an increment of cell apoptosis by the combination of simvastatin and LBH589 in a mouse xenograft model. These data suggest that simvastatin is a novel Hsp90 inhibitor to disrupt the formation of the K292-acetylated Hsp90/Cdc37 complex in triple-negative breast cancer cells. The combination of simvastatin with LBH589 could be used as a novel therapeutic strategy for triple-negative breast cancer.

Honokiol, a bioactive compound isolated from the cone and bark of Magnolia officinalis, has been shown to have various activities including inhibition of the growth of Candida albicans. We investigated the roles of the Hsp90-calcineurin pathway in the antifungal activity of honokiol. The pharmacologic tool was employed to evaluate the effects of Hsp90 and calcineurin in the antifungal activity of honokiol. We also evaluated the protective effects of the calcineurin inhibitor cyclosporin A (CsA) on honokiol-induced mitochondrial dysfunction by the fluorescence staining method. The Hsp90 inhibitor potentiated the antifungal activity of honokiol. A C. albicans strain with the calcineurin gene deleted displayed enhanced sensitivity to honokiol. However, co-treatment with calcineurin inhibitor CsA attenuated the cytotoxic activity of honokiol due to the protective effect on mitochondria. Our results provide insight into the action mechanism of honokiol.

Fibronectin (FN) plays a major role in the stability and organization of the extracellular matrix (ECM). We have previously demonstrated that FN interacts directly with Hsp90, as well as showing that the Hsp90 inhibitor novobiocin results in FN turnover via a receptor mediated process. However, the receptor involved has not been previously identified. LRP1 is a ubiquitous receptor responsible for the internalisation of numerous ligands that binds both Hsp90 and FN, and therefore we investigated whether LRP1 was involved in novobiocin-mediated FN turnover. FN, LRP1 and Hsp90 could be isolated in a common complex, and inhibition of Hsp90 by novobiocin increased the colocalisation of FN and LRP1. Novobiocin induced an increase (at low concentrations) followed by a loss of FN that was primarily derived from extracellular matrix-associated FN and led to a concomitant increase in intracellular FN. The effect of novobiocin was specific to LRP1-expressing cells and could be recapitulated by an LRP1 blocking antibody and the allosteric C-terminal Hsp90 inhibitor SM253, but not the N-terminal inhibitor geldanamycin. Together these data suggest that LRP1 is required for FN turnover in response to Hsp90 inhibition by novobiocin, which may have unintended physiological consequences in contexts where C-terminal Hsp90 inhibition is to be used therapeutically.

Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer with increases in new cases being reported annually. Histopathologists have identified hepatic steatosis as a characteristic of a broad range of chronic liver diseases that are associated with the onset and development of HCC. In this context, epigenetic modifications may serve as precancerous factors predisposing normal cells to the initiation of carcinogenesis. This study demonstrated that hepatic tumorigenesis and differentiated adipocytes may modulate both global histone deacetylase (HDAC) expression and specific class I HDAC genes in the tumour microenvironment. The novel class I HDAC inhibitor Resminostat was shown to reduce the proliferation of HCC cells along with its specificity in targeting class I HDACs and oncogenes. The combined effect of Resminostat with several pharmaceutical agents such as Sorafenib, Cisplatin and Doxorubicin was also demonstrated. The inhibition of heat shock protein 90 (HSP90) has been demonstrated as a potential therapeutic option for HCC. In line with this, the specific HSP90 inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17-AAG) was selected and it was found that the combination of Resminostat and 17-AAG may provide a "smart" clinical strategy for HCC patients by targeting cellular communication within the tumour microenvironment. This study provides an insight into the use of Resminostat as an epigenetic based therapeutic for HCC along with other pharmaceutical options, in particular by targeting the cell-to-cell communication that occurs between hepatoma and adipocytes.

Heat shock protein 90 (Hsp90) is an evolutionary preserved molecular chaperone which mediates many cellular processes such as cell transformation, proliferation, and survival in normal and stress conditions. Hsp90 plays an important role in folding, maturation, stabilization and activation of Hsp90 client proteins which all contribute to the development, and proliferation of cancer as well as other inflammatory diseases. Functional inhibition of Hsp90 can have a massive effect on various oncogenic and inflammatory pathways, and will result in the degradation of their client proteins. This turns it into an interesting target in the treatment of different malignancies. 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) as a semi-synthetic derivative of geldanamycin, has several advantages over 17-Allylamino-17-demethoxygeldanamycin (17-AAG) such as higher water solubility, good bioavailability, reduced metabolism, and greater anti-tumour capability. 17-DMAG binds to the Hsp90, and inhibits its function which eventually results in the degradation of Hsp90 client proteins. Here, we reviewed the pre-clinical data and clinical trial data on 17-DMAG as a single agent, in combination with other agents and loaded on nanomaterials in various cancers and inflammatory diseases.

Heat shock protein 90 (HSP90) inhibitors suppressed MDM4 functions which mediated p53 ubiquitination, and blocked a chaperon function which influenced expression of the client proteins. We examined cytotoxic effects of the inhibitors, 17-allylamino-17-demetheoxygeldanamycin (17-AAG) and 17-dimethylaminoethylamino-17-demethoxy-geldanamycin (17-DMAG), on mesothelioma and investigated combinatory effects of the inhibitors and adenoviruses expressing the wild-type p53 gene (Ad-p53). A majority of mesothelioma lacks p14 and p16 expression, which leads to defective p53 pathway despite bearing the wild-type p53 genotype. The HSP90 inhibitors up-regulated endogenous wild-type p53 expression and induced cell death. Furthermore, the inhibitors increased the endogenous p53 levels that were induced by cisplatin. Nevertheless, the HSP90 inhibitors suppressed Ad-p53-induced exogenous p53 expression primarily at a posttranscriptional level and inhibited the Ad-p53-mediated cell death. HSP90 inhibitors suppressed ubiquitination processes which were involved in p53 degradation, but a proteasome inhibitor, MG-132, prevented the HSP90 inhibitors-induced p53 down-regulation. In contrast, an inhibitor for HSP70 with a chaperon function, pifithrin-μ, did not produce the p53 down-regulation. The HSP90 inhibitors did not suppress expression of Ad receptor molecules but rather increased expression of green fluorescence protein transduced by the same Ad vector. These data collectively indicated that an HSP90 inhibitor possessed a divalent action on p53 expression, as an activator for endogenous wild-type p53 through inhibited ubiquitination and a negative regulator of exogenously over-expressed p53 through the proteasome pathway.

Although ErbB2-targeted therapeutics have significantly improved ErbB2+ breast cancer patient outcomes, therapeutic resistance remains a significant challenge. Therefore, the development of novel ErbB2-targeting strategies is necessary. Importantly, ErbB2 is a sensitive client protein of heat shock protein 90 (HSP90), which regulates client protein folding, maturation, and stabilization. HSP90 inhibition provides an alternative therapeutic strategy for ErbB2-targeted degradation. In particular, ganetespib, a novel HSP90 inhibitor, is a promising agent for ErbB2+ cancers. Nevertheless, the anti-cancer efficacy and clinical application of ganetespib for ErbB2+ breast cancer is largely unknown. In our study, we examined the anti-cancer effects of ganetespib on ErbB2+ BT474 and SKBR3 breast cancer cells, and isogenic paired cancer cell lines with lentivirus-mediated ErbB2 overexpression. Ganetespib potently inhibited cell proliferation, cell cycle progression, survival, and activation/phosphorylation of ErbB2 and key downstream effectors in ErbB2+ breast cancer cells. Moreover, ganetespib decreased the total protein levels of HSP90 client proteins and reduced ErbB2 protein half-life. ErbB2-overexpressing cancer cells were also more sensitive to ganetespib-mediated growth inhibition than parental cells. Ganetespib also strikingly potentiated the inhibitory effects of lapatinib in BT474 and SKBR3 cells. Ultimately, our results support the application of ganetespib-mediated HSP90 inhibition as a promising therapeutic strategy for ErbB2+ breast cancer.

Esophageal carcinoma is aggressive in nature and its prognosis is largely dependent on the degree of invasion. Histone deacetylase 6 (HDAC6), as the most unique member of HDACs family, has the positive activity to promote initiation and progression of various cancers via targeting multiple non‐histone proteins in cytoplasm. In this study, we found that HDAC6 was over‐expressed in three esophageal cancer cell lines (KYSE140, KYSE170, KYSE180) when compared to non‐carcinoma esophageal epithelial cell HEEC‐1. Then two HDAC6 specific siRNAs and HDAC6 inhibitor tubastatin A greatly suppressed KYSE140 and KYSE180 cells proliferation and migration, and the inhibition of cell motility was accompanied by elevated acetylation of α‐tubulin, a target of HDAC6. Consistently, the microtubulin skeleton was stabilized after HDAC6 knockdown or inhibition. In addition, acetylation status of HSP90, another HDAC6 target, was also increased towards HDAC6 knockdown or inhibition by co‐immunoprecipitation assay. Besides, co‐treatment of HSP90 inhibitor (PU‐H71) and HDAC6 inhibitor (tubastatin A) induced a stronger cell migration inhibition compared to administration of either drug alone. Furthermore, cell proliferation of KYSE140 and KYSE180 were also compromised in response to combination of HDAC6 and HSP90 inhibitors. Additionally, co‐administration of HSP90 inhibitor and HDAC6 inhibitor strongly inhibited tumor growth in vivo. Taken together, our results indicated that HDAC6 is a promising target by inhibiting HSP90 function in ESCC.

Introduction: In this phase 2 study, we evaluated the activity of AUY922 in pretreated patients with stage IV NSCLC. Methods: Patients with advanced NSCLC were divided into molecularly defined strata based on mutations in the EGFR gene, the ALK receptor tyrosine kinase gene (ALK), the KRAS gene, or the wild type of all three. All patients must have received more than two prior lines of therapy, except for those in a fifth stratum for a less pretreated EGFR cohort (EGFR= 40%) all-causality adverse events were diarrhea, nausea, and decreased appetite. Visual-related disorders were reported in 79.7% of patients (most were grade 1/2). Thirty-five patients (22.9%) reported night blindness. Conclusion: AUY922 is active in patients with NSCLC, particularly among patients with ALK rearrangements and EGFR mutations. (C) 2017 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.

A series of 1-aroylindoline-hydroxamic acids have been synthesized in the present study. The results of the biological evaluation led to the identification of compound 12 as dual HDAC6/HSP90 inhibitor. Compound 12 displayed striking inhibitory effects towards the HDAC6 isoform and HSP 90 protein with IC50 values of 1.15 nM (HDAC6) and 46.3 nM (HSP90). Compound 12 also exhibited 113, 139 and 246 fold higher selectivity for HDAC6 over HDAC 1, HDAC 3 and HDAC 8 isoforms and was endowed with significant cytotoxic effects with GI50 values ranging 1.04–1.61 μM against lung A549, colorectal HCT116, leukemia HL60, and EGFR T790M mutant lung H1975 cell lines. Another interesting finding of the study was substantial cytotoxic effects of compounds particularly against lung H1975 (NSCLC) cell lines with IC50 = 0.26 μM which may be mediated through HSP90 inhibition. Compound 8 as such was devoid of HDAC inhibitory activity.

The BRAFV600E mutation results in the constitutive activation of downstream mitogen activated protein kinase (MAPK) pathway that promotes tumor growth. Recently, molecular targeted therapy using BRAF/MEK inhibitor has been reported for BRAFV600E mutant high-grade glioma, but the therapeutic effect is limited by the emergence of drug resistance. Herein, we established paired BRAFV600E mutant glioblastoma (GBM) patient-derived xenograft (PDX) models, which were derived from tumors at prior to and recurrence after molecular targeted therapy. These PDX models were found to extensively recapitulate the histology, genetic abnormalities, and even the clinical course of the patients. Furthermore, BRAF/MEK inhibitor gradually caused resistance in cell lines derived from specimens that initially responded to molecular targeted therapy. In this study, genomic and epigenomic changes had little effect on the resistance mechanism. On the other hand, we found that hyperactivation of the MAPK pathway through c-Raf and the AKT/mTOR pathway primarily caused resistance to molecular targeted therapy in BRAFV600E mutant GBM. Through a high throughput drug screening, we find that HSP90 inhibitor with BRAF/MEK inhibitor coordinately deactivates MAPK pathway and AKT/mTOR pathway, and mediates potent toxicity in vitro and in vivo in refractory and acquired resistant models. These findings support that this therapeutic approach can overcome the limitation of current molecular targeted therapy in BRAFV600E mutant GBM.

Fluorescent imaging has been widely used in the detection of diseases, especially cancer. Among them, biomarker-specific ligand-based imaging strategies have attracted much attention, due to their improved image contrast and diagnostic accuracy. In this work, an HSP90 inhibitor based fluorescent probe (Probe 1) was synthesized possessing the potential of selective tumor targeting, which was constructed with an HSP90 inhibitor and 4-hydroxy-1,8-naphthalimide derivative. Probe 2, a resorcinol-blocked probe was also synthesized as a control, which prevented binding with the HSP90 ATP binding pocket. Biological studies showed that Probe 1 could be selectively accumulated by tumor cells, while Probe 2 not, suggesting that Probe 1 is a promising targeted tumor imaging agent for early diagnosis.

Currently available SARS-CoV-2 therapeutics are targeted towards moderately to severely ill patients and require intravenous infusions, with limited options for exposed or infected patients with no or mild symptoms. While vaccines have demonstrated protective efficacy, vaccine hesitancy and logistical distribution challenges will delay their ability to end the pandemic. Hence, there is a need for rapidly translatable, easy-to-administer-therapeutics, that can prevent SARS-CoV-2 disease progression, when administered in the early stages of infection. We demonstrate that an orally bioavailable Hsp90 inhibitor, SNX-5422, currently in clinical trials as an anti-cancer therapeutic, inhibits SARS-CoV-2 replication in vitro at a high selectivity index. SNX-5422 treatment of human primary airway epithelial cells dampened expression of inflammatory pathways previously associated with poor SARS-CoV-2 disease outcomes. Additionally, SNX-5422 interrupted expression of host factors demonstrated to be crucial for SARS-CoV-2 replication. Development of SNX-5422 as SARS-CoV-2-early-therapy will dampen disease severity, resulting in better clinical outcomes and reduced hospitalizations., Graphical Abstract