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Objective: Hepatocellular carcinoma (HCC), the main type of liver cancer, has a high morbidity and mortality, and a poor prognosis. RNA helicase DDX5, which acts as a transcriptional co-regulator, is overexpressed in most malignant tumors and promotes cancer cell growth. Heat shock protein 90 (HSP90) is an important molecular chaperone in the conformational maturation and stabilization of numerous proteins involved in cell growth or survival. Methods: DDX5 mRNA and protein expression in surgically resected HCC tissues from 24 Asian patients were detected by quantitative real-time PCR and Western blot, respectively. The interaction of DDX5-HSP90 was determined by molecular docking, immunoprecipitation, and laser scanning confocal microscopy. The autophagy signal was detected by Western blot. The cell functions and signaling pathways of DDX5 were determined in 2 HCC cell lines. Two different murine HCC xenograft models were used to determine the function of DDX5 and the therapeutic effect of an HSP90 inhibitor. Results: HSP90 interacted directly with DDX5 and inhibited DDX5 protein degradation in the AMPK/ULK1-regulated autophagy pathway. The subsequent accumulation of DDX5 protein induced the malignant phenotype of HCC by activating the β-catenin signaling pathway. The silencing of DDX5 or treatment with HSP90 inhibitor both blocked in vivo tumor growth in a murine HCC xenograft model. High levels of HSP90 and DDX5 protein were associated with poor prognoses. Conclusions: HSP90 interacted with DDX5 protein and subsequently protected DDX5 protein from AMPK/ULK1-regulated autophagic degradation. DDX5 and HSP90 are therefore potential therapeutic targets for HCC.

Kinesin-like calmodulin-binding protein (KCBP) is a unique kinesin with half kinesin and half myosin, with kinesin motor domain at C-terminus and myosin tail homology region 4 (MyTH4) and band 4.1, ezrin, radixin, moesin (FERM) domains at N-terminus. The special structure endows KCBP multi-intracellular functions, including cell division, trichome morphogenesis in plants, and flagellar function in algae. However, little is known about the molecular mechanism underlying these functions. Here, we identified a molecular chaperone Hsp90 as a novel binding partner with KCBP in Dunaliella salina using a yeast two-hybrid screen. Further analysis showed that Hsp90 interacted with both the N-terminal and C-terminal of DsKCBP. Since Hsp90 was involved in the stability and proteolytic turnover of numerous proteins, whether Hsp90 regulated the degradation of DsKCBP was investigated. Our results showed that both Hsp90 and DsKCBP presented in the purified proteasome, and the interaction of DsKCBP-Hsp90 was inhibited upon Hsp90 inhibitor geldanamycin treatment. The level of DsKCBP proteins was diminished remarkably indicating that the disassociation of DsKCBP from Hsp90 accelerated the degradation of the former. Furthermore, immunofluorescence results showed that the localization of DsKCBP at basal body and flagella was disappeared by Hsp90 inhibition. The increased mRNA level of DsKCBP during flagellar assembly was not obvious by geldanamycin treatment. These data provided evidence that Hsp90 protected DsKCBP from degradation by proteasome and was involved in the role of DsKCBP in flagellar assembly.

Heat shock protein 90 (HSP90) is involved in plant growth and various stress responses via regulating protein homeostasis. Autophagy keeps cellular homeostasis by recycling the components of cellular cytoplasmic constituents. Although they have similar effects on cellular protein homeostasis, the direct association between HSP90 and autophagy signaling remains unclear in plants, especially in tropical crops. In this study, the correlation between HSP90 and autophagy signaling was systematically analyzed by protein-protein interaction in cassava, one of the most important economy fruit in tropic. In addition, their effects on plant disease response and underlying mechanisms in cassava were investigated by functional genomics and genetic phenotype assay. The potential MeHSP90.9-MeSGT1-MeRAR1 chaperone complex interacts with MeATGs and subsequently triggers autophagy signaling, conferring improved disease resistance to cassava bacterial blight (CBB). On the contrary, HSP90 inhibitor and autophagy inhibitor decreased disease resistance against CBB in cassava, and autophagy may be involved in the potential MeHSP90.9-MeSGT1-MeRAR1 chaperone complex-mediated multiple immune responses. This study highlights the precise modulation of autophagy signaling by potential MeHSP90.9-MeSGT1-MeRAR1 chaperone complex in autophagy-mediated disease resistance to CBB.

Background Heat shock protein 90 (HSP90) is a molecular chaperone that is responsible for the conformational maintenance of several client proteins that play important roles in DNA damage repair, apoptosis following radiation, and resistance to radiation therapy. DS-2248 (tricyclic pyrazolopyrimidine derivative) is a newly-developed, orally available inhibitor of HSP90 with low adverse effects. We investigated the combined effects of radiation and DS-2248 in vitro and in vivo. Methods SCCVII squamous cell carcinoma cells and tumors transplanted in C3H/HeN mice were used. In vitro combined effects of X-ray radiation and DS-2248 were investigated using a colony assay. Phosphorylated histone H2AX (γH2AX) was quantified after 2-Gy irradiation with or without 24-hour pretreatment with DS-2248. The mice bearing SCCVII tumors received oral DS-2248 10 times over 2 weeks and received local irradiation with doses of 1, 2, 3, and 4 Gy delivered 6 times over 2 weeks. Then, tumor volumes were measured. Results Radiation plus pretreatment with 50 nM DS-2248 for 24 hours produced synergistic effects on SCCVII cells. γH2AX foci persisted after radiation for longer periods (6 and 24 hours) in DS-2248-treated cells than in control cells. In vivo, the combined effects appeared to be additive when 5 or 10 mg/kg DS-2248 was combined with total radiation doses of 6–18 Gy, but the effect was considered supra-additive when 15 mg/kg of DS-2248 was combined with a total dose of 24 Gy. Conclusions The combined effects of DS-2248 and radiation were additive at low drug and radiation doses, but may have been supra-additive at higher doses. Inhibition of slow repair of DNA double strand breaks (i.e., homologous recombination) was considered to contribute to this combined effect.

The heat shock protein 90 inhibitor, luminespib, has demonstrated potent preclinical activity against numerous cancers. However, clinical translation has been impeded by dose-limiting toxicities that have necessitated dosing schedules which have reduced therapeutic efficacy. As such, luminespib is a prime candidate for reformulation using advanced drug delivery strategies that improve tumor delivery efficiency and limit off-target side effects. Specifically, thermosensitive liposomes are proposed as a drug delivery strategy capable of delivering high concentrations of drug to the tumor in combination with other chemotherapeutic molecules. Indeed, this work establishes that luminespib exhibits synergistic activity in lung cancer in combination with standard of care drugs such as cisplatin and vinorelbine. While our research team has previously developed thermosensitive liposomes containing cisplatin or vinorelbine, this work presents the first liposomal formulation of luminespib. The physico-chemical properties and heat-triggered release of the formulation were characterized. Cytotoxicity assays were used to determine the optimal drug ratios for treatment of luminespib in combination with cisplatin or vinorelbine in non-small cell lung cancer cells. The formulation and drug combination work presented in this paper offer the potential for resuscitation of the clinical prospects of a promising anticancer agent.

Heat shock protein 90 (Hsp90) is a key chaperone that is abnormally expressed in cancer cells, and therefore, designing novel compounds to inhibit chaperone activities of the Hsp90 is a promising therapeutic approach for cancer drug discovery. Debio-0932 is a second-generation Hsp90 inhibitor that exhibited promising anticancer activity against a wide variety of cancer types with a strong binding affinity for Hsp90 and high oral bioavailability. Anticancer activities of the Debio-0932 were tested in MCF-7 and MDA-MB-231 cell lines. Molecular docking results indicated that Debio-0932 was selectively bound to the ATP binding pocket of the Hsp90 with an estimated free energy of binding − 7.24 kcal/mol. Antiproliferative activity of Debio-0932 was determined by XTT assay and Debio-0932 exhibited a cytotoxic effect on MCF-7 and MDA-MB-231 cells in a time and dose-depended manner. Apoptosis inducer role of Debio-0932 was evaluated in MCF-7 and MDA-MB-231 cells with fluorometric apoptosis/necrosis detection kit. Treatment with Debio-0932 stimulated apoptosis in both breast cancer cell lines. mRNA and protein expression levels of Bax, Bcl-2 and Casp-9 were determined in MCF-7 and MDA-MB-231 cells by RT-PCR and Western blotting respectively. Debio-0932 stimulated the down-regulation of anti-apoptotic protein Bcl-2 and the up-regulation of apoptotic protein Bax and cleavage of Casp-9 in cancer cells. Moreover, the anti-invasive potential of Debio-0932 was evaluated in endothelial cells (HUVEC) by wound-healing assay. Debio-0932 decreased the migration of HUVEC cells as compared to the control group. These results indicate that Debio-0932 is a promising compound to treat triple-negative breast cancer and hormone receptor-positive breast cancer, and their metastases.

Chenwei Pan,1,* Tiaotiao Zhang,2,3,* Shaoxun Li,1,3,* Zhihua Xu,2,3 Binhui Pan,2,3 Sheng Xu,2,3 Shuanghong Jin,1,3 Guangrong Lu,2 Shouxing Yang,2 Zhanxiong Xue,2 Ping Chen,4 Xian Shen,5 Fangyan Wang,6 Changlong Xu2,7 1Department of Infectious Disease, Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, 325000, People’s Republic of China; 2Department of Gastroenterology, Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, 325000, People’s Republic of China; 3Second School of Medicine, Wenzhou Medical University, Wenzhou, Zhejiang, 325000, People’s Republic of China; 4Department of Infectious Disease, Shulan Hospital, Hangzhou, Zhejiang, 310012, People’s Republic of China; 5Department of Gastrointestinal Surgery, Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, 325000, People’s Republic of China; 6Department of Pathophysiology, School of Basic Medicine Science, Wenzhou Medical University, Wenzhou, Zhejiang, 325000, People’s Republic of China; 7Center for Diagnostics and Therapeutics, Institute for Biomedical Sciences, Georgia State University, Atlanta, GA, 30302, USA*These authors contributed equally to this workCorrespondence: Fangyan Wang; Changlong Xu Email wzyxywfy@126.com; xchlong@163.comBackground: As a therapeutic target for cancer treatment, HSP90 has been explored extensively. However, the significant side effects of the HSP90 inhibitor 17AAG have limited its clinical use.Methods: In this study, we used hyaluronic acid (HA)–decorated DOTAP–PLGA hybrid nanoparticles (HA-DOTAP-PLGA NPs) as 17AAG-delivery carriers for targeted colon cancer therapy.Results: Different methods were used to characterize the successful fabrication of these hybrid PLGA NPs. Our results demonstrated that internalization of HA-NPs in colon cancer cells was governed by CD44receptor–mediated endocytosis. Annexin V–propidium iodide staining experiments revealed that cell apoptosis induced by HA-NPs-17AAG in colon cancer cells was more efficient than free 17AAG. In two animal models used to screen anticancer efficacy (Luc-HT29 subcutaneous xenograft and AOM/DSS-induced orthotopic tumor model), HA-NPs-17AAG significantly inhibited xenograft and orthotopic tumor growth, demonstrating HA-NPs-17AAG had much better therapeutic efficiency than free 17AAG. It is worth noting that great biocompatibility of HA-DOTAP-PLGA NPs was observed both in vitro and in vivo.Conclusion: Our research offers a preclinical proof of concept for colon cancer therapy with DOTAP-PLGA NPs as a creative drug-delivery system.Keywords: cationic hybrid nanoparticles, hyaluronic acid, HA, HSP90 inhibitor, colon cancer, targeted delivery system

Background Patients with non‐small cell lung cancer (NSCLC) harboring activating EGFR mutations are sensitive to epidermal growth factor receptor‐tyrosine kinase inhibitors (EGFR‐TKIs) but inevitably develop resistance to the inhibitors mostly through acquisition of the secondary T790M mutation. Although third‐generation EGFR‐TKIs overcome this resistance by selectively inhibiting EGFR with EGFR‐TKI‐sensitizing and T790M mutations, acquired resistance to third‐generation EGFR‐TKIs invariably develops. Methods Next‐generation sequencing (NGS) and fluorescence in situ hybridization (FISH) analysis were performed in an EGFR T790M‐mutated NSCLC patient who had progressed after a third‐generation EGFR‐TKI, TAS‐121. EGFR‐mutated cell lines were subjected to a cell proliferation assay and western blotting analysis with EGFR‐TKIs and a heat shock protein 90 (HSP90) inhibitor. Results NGS and FISH analysis revealed EGFR amplification in the resistant cancer cells. While EGFR L858R/T90M‐mutated cell line was sensitive to osimertinib or TAS‐121 in vitro, EGFR‐overexpressing cell lines displayed resistance to these EGFR‐TKIs. Western blot analysis showed that EGFR phosphorylation and overexpression of EGFR in cell lines was not suppressed by third‐generation EGFR‐TKIs. In contrast, an HSP90 inhibitor reduced total and phosphorylated EGFR and inhibited the proliferation of resistant cell lines. Conclusions EGFR amplification confers resistance to third‐generation EGFR‐TKIs which can be overcome by HSP90 inhibition. The results provide a preclinical rationale for the use of HSP90 inhibitors to overcome EGFR amplification‐mediated resistance., Third‐generation EGFR‐TKIs overcome EGFR T790M‐meditated resistance. However, amplified EGFR confers resistance to third‐generation EGFR‐TKIs. Inhibiting HSP90, a chaperon for EGFR, suppresses the proliferation of EGFR‐amplified cells by degrading EGFR in vitro, providing a preclinical rationale for the use of HSP90 inhibitors to overcome the EGFR amplification‐meditated resistance.

Treatment relapse due to clonal evolution was shown to be an independent factor for poor prognosis in advanced stages of chronic myeloid leukemia. Overcoming secondary resistance arising due to clonal evolution is still an unmet need and lack of adequate pre-clinical models hampers the identification of underlying mechanisms and testing of alternate treatment strategies. The current study thus aimed to create cellular models to study molecular mechanisms underlying clonal evolution and identify strategies to overcome the secondary drug resistance. Analysis of cell lines derived from three independent cell-based screens revealed the co-evolution specifically of imatinib and HSP90 inhibitor (HSP90i) resistances despite their exposure to a single inhibitor alone. Molecular and biochemical characterization of these cell lines revealed additional cytogenetic abnormalities, differential activation of pro-survival signaling molecules and over expression of ABL kinase and HSP90 genes. Importantly, all the imatinib-HSP90i dual resistant cell lines remained sensitive to sorafenib and vorinostat suggesting their utility in treating patients who relapse upon imatinib treatment due to clonal evolution. In addition, we cite similar examples of dual resistance towards various kinase inhibitors and HSP90i in some cell lines that represent solid cancers suggesting co-evolution leading to secondary drug resistance as a pan-cancer phenomenon. Taken together, our results suggest the efficacy of HSP90i in overcoming drug resistance caused by point mutations in the target kinase but not in cases of clonal evolution.

The use of targeted inhibitors has shown promise as an effective approach in cancer therapy. However, targeted therapies based only on one drug, such as 17-allylamino-17-demethoxygeldanamycin (17-AAG), have limited success, partly because cancer cells engage alternate pathways for survival and proliferation. In the present study, we evaluated whether dehydroxymethylepoxyquinomicin (DHMEQ), a nuclear factor ?B (NF-?B) inhibitor, can enhance the antitumor activities of 17-AAG, a 90 kDa heat shock protein (Hsp90) inhibitor, in the anaplastic thyroid cancer cell line KTC2. We examined the effect of combined drug treatment vs single drug treatment on cell survival. Isobologram analysis was performed to distinguish the additive vs synergistic effects of the drug combination. Western blotting was performed to investigate apoptosis markers: caspase 3, poly(ADP-ribose) polymerase-one (PARP-1), B-cell lymphoma-extra large (Bcl-XL), X-linked inhibitor of apoptosis (XIAP) and cellular inhibitor of apoptosis 2 (cIAP-2). Compared to monotherapy, the combined treatment enhanced growth-inhibitory effects in a synergistic manner and strongly potentiated apoptosis. These results demonstrate the first in vitro evidence that a combination of Hsp90 and NF-?B inhibitors is a more effective modality for inhibiting cell proliferation and survival in anaplastic thyroid carcinoma cells than either agent alone, warranting further investigations.

BackgroundPhototherapy is a promising strategy for cancer therapy by reactive oxygen species (ROS) of photodynamic therapy (PDT) and hyperthermia of photothermal therapy (PTT). However, the therapeutic efficacy was restricted by tumor hypoxia and thermal resistance of increased expression of heat shock protein (Hsp). In this study, we developed albumin nanoparticles to combine hypoxia relief and heat shock protein inhibition to overcome these limitations for phototherapy enhancement.ResultsNear-infrared photosensitizer (IR780) and gambogic acid (GA, Hsp90 inhibitor) were encapsulated into albumin nanoparticles via hydrophobic interaction, which was further deposited MnO2on the surface to form IGM nanoparticles. Both in vitro and in vivo studies demonstrated that IGM could catalyze overexpress of hydrogen peroxide to relive hypoxic tumor microenvironment. With near infrared irradiation, the ROS generation was significantly increase for PDT enhancement. In addition, the release of GA was promoted by irradiation to bind with Hsp90, which could reduce cell tolerance to heat for PTT enhancement. As a result, IGM could achieve better antitumor efficacy with enhanced PDT and PTT.ConclusionThis study develops a facile approach to co-deliver IR780 and GA with self-assembled albumin nanoparticles, which could relive hypoxia and suppress Hsp for clinical application of cancer phototherapy.

Purpose: Non-Small-Cell Lung Cancer (NSCLC) has gained resistance to common chemo- and radiotherapy due to the oncogenic K-RAS mutations. In this work, lactonic sophorolipids (LSL), a constituent of natural sophorolipids known to inhibit histone deacetylase (HDAC) activity, is used to evaluate its potential anticancer property for the treatment of NSCLC. In addition, ganetespib (GT), a Hsp90 inhibitor, is used for its known antitumor activity in several K-RAS mutant NSCLC cells. We propose, a functional anti-oxidant nanomedicine composed of nanoceria (NC) encapsulated with two-drug cocktail LSL and GT for the assessment of therapeutic efficacy of LSL and targeted combination therapy of NSCLC. NC is an excellent redox platform specifically used to supplement the therapeutic potency of these drugs to target both HDAC inhibition and Hsp90 signaling pathways in NSCLC. Methods: Polyacrylic acid-coated nanoceria (PNC) was formulated and folic acid was conjugated on the surface of PNC using “click” chemistry to target NSCLC and to minimize adverse side effects. Solvent diffusion method was used for the encapsulation of individual drugs and co-encapsulation of drug-cocktail along with an optical dye DiI for diagnosis. We hypothesized that the therapeutic efficacy of LSL will be synergistically accelerated by the inhibition of Hsp90 mechanism of GT and redox activity of NC. Results: For the targeted therapy of NSCLC, A549 cells were used and Chinese hamster ovary (CHO) cells were used as healthy control cells. Results showed more than 40% cells were dead within 24 h when treated with LSL nanodrug. When combined with GT, enhanced ROS signals were detected and more than 80% reduction in cell viability was recorded within 24 h of incubation. Treatments with NC without any drug showed minimal toxicity. Migration assays indicate that the highly metastatic nature of NSCLC is successfully restricted by this combination approach. To validate the effectiveness of this combination therapy various cell-based assays including detection of apoptosis, necrosis and HDAC inhibition of LSL were performed. Conclusion: Functional nanoceria with drug-cocktail LSL and GT is successfully developed for the targeted treatment of undruggable NSCLC. The fluorescence modality helps monitoring the drugs delivery. Results demonstrate the potential therapeutic efficacy of LSL, which is synergistically accelerated by the Hsp90 inhibition mechanism of GT and redox activity of NC.

Many viruses utilize molecular chaperone heat shock protein 90 (Hsp90) for protein folding and stabilization, however, the role of Hsp90 in herpesvirus lifecycle is obscure. Here, we provide evidence that Hsp90 participates in pseudorabies virus (PRV) replication. Viral growth kinetics assays show that Hsp90 inhibitor geldanamycin (GA) abrogates PRV replication at the post-penetration step. Transmission electron microscopy demonstrates that dysfunction of Hsp90 diminishes the quantity of PRV nucleocapsids. Overexpression and knockdown of Hsp90 suggest that de novo Hsp90 is involved in PRV replication. Mechanismly, dysfunction of Hsp90 inhibits PRV major capsid protein VP5 expression. Co-immunoprecipitation and indirect immunofluorescence assays indicate that Hsp90 interacts with VP5. Interestingly, Hsp70, a collaborator of Hsp90, also interacts with VP5, but doesn't affect PRV growth. Finally, inhibition of Hsp90 results in PRV VP5 degradation in a proteasome-dependent manner. Collectively, our data suggest that Hsp90 contributes to PRV virion assembly and replication via stabilization of VP5.

Drug-induced resistance, or tolerance, is an emerging yet poorly understood failure of anticancer therapy. The interplay between drug-tolerant cancer cells and innate immunity within the tumor, the consequence on tumor growth, and therapeutic strategies to address these challenges remain undescribed. Here, we elucidate the role of taxane-induced resistance on natural killer (NK) cell tumor immunity in triple-negative breast cancer (TNBC) and the design of spatiotemporally controlled nanomedicines, which boost therapeutic efficacy and invigorate “disabled” NK cells. Drug tolerance limited NK cell immune surveillance via drug-induced depletion of the NK-activating ligand receptor axis, NK group 2 member D, and MHC class I polypeptide-related sequence A, B. Systems biology supported by empirical evidence revealed the heat shock protein 90 (Hsp90) simultaneously controls immune surveillance and persistence of drug-treated tumor cells. On the basis of this evidence, we engineered a “chimeric” nanotherapeutic tool comprising taxanes and a cholesterol-tethered Hsp90 inhibitor, radicicol, which targets the tumor, reduces tolerance, and optimally reprimes NK cells via prolonged induction of NK-activating ligand receptors via temporal control of drug release in vitro and in vivo. A human ex vivo TNBC model confirmed the importance of NK cells in drug-induced death under pressure of clinically approved agents. These findings highlight a convergence between drug-induced resistance, the tumor immune contexture, and engineered approaches that consider the tumor and microenvironment to improve the success of combinatorial therapy. Significance: This study uncovers a molecular mechanism linking drug-induced resistance and tumor immunity and provides novel engineered solutions that target these mechanisms in the tumor and improve immunity, thus mitigating off-target effects.

Summary Excessive and dysregulated inflammation is known to contribute to disease progression. HSP90 is an intracellular chaperone known to regulate inflammatory processes including the NLRP3 inflammasome and secretion of the pro‐inflammatory cytokine interleukin(IL)‐1β. Here, primarily using an in vitro inflammasome ASC speck assay, and an in vivo model of murine peritonitis, we tested the utility of HSP90 inhibitors as anti‐inflammatory molecules. We report that the HSP90 inhibitor EC144 effectively inhibited inflammatory processes including priming and activation of NLRP3 in vitro and in vivo. A specific inhibitor of the β HSP90 isoform was ineffective suggesting the importance of the α isoform in inflammatory signalling. EC144 inhibited IL‐1β and IL‐6 in vivo when administered orally, and was brain‐penetrant. These data suggest that HSP90 inhibitors may be useful for targeting inflammation in diverse diseases that are worsened by the presence of inflammation., HSP90 inhibitors inhibit priming and activation of the NLRP3 inflammasome.

Purpose:Treatment approaches using Hsp90 inhibitors at their maximum tolerated doses (MTDs) have not produced selective tumor toxicity. Inhibition of Hsp90 activity causes degradation of client proteins including those involved in recognizing and repairing DNA lesions. We hypothesized that if DNA repair proteins were degraded by concentrations of an Hsp90 inhibitor below those required to cause nonspecific cytotoxicity, significant tumor-selective radiosensitization might be achieved.Experimental Design:Tandem mass tagged-mass spectrometry was performed to determine the effect of a subcytotoxic concentration of the Hsp90 inhibitor, AT13387 (onalespib), on global protein abundance. The effect of AT13387 on in vitro radiosensitization was assessed using a clonogenic assay. Pharmacokinetics profiling was performed in mice bearing xenografts. Finally, the effect of low-dose AT13387 on the radiosensitization of three tumor models was assessed.Results:A subcytotoxic concentration of AT13387 reduced levels of DNA repair proteins, without affecting the majority of Hsp90 clients. The pharmacokinetics study using one-third of the MTD showed 40-fold higher levels of AT13387 in tumors compared with plasma. This low dose enhanced Hsp70 expression in peripheral blood mononuclear cells (PBMCs), which is a biomarker of Hsp90 inhibition. Low dose monotherapy was ineffective, but when combined with radiotherapy, produced significant tumor growth inhibition.Conclusions:This study shows that a significant therapeutic ratio can be achieved by a low dose of Hsp90 inhibitor in combination with radiotherapy. Hsp90 inhibition, even at a low dose, can be monitored by measuring Hsp70 expression in PBMCs in human studies.

HSP90 is a molecular chaperone required for stabilization and activation of hundreds of client proteins, including many known oncoproteins. AUY922 (luminespib), a new-generation HSP90 inhibitor, exhibits potent preclinical efficacy against several cancer types including prostate cancer. However, clinical use of HSP90 inhibitors for prostate cancer has been limited by toxicity and treatment resistance. Here, we aimed to design an effective combinatorial therapeutic regimen that utilizes subtoxic doses of AUY922, by identifying potential survival pathways induced by AUY922 in clinical prostate tumors. We conducted a proteomic analysis of 30 patient-derived explants (PDE) cultured in the absence and presence of AUY922, using quantitative mass spectrometry. AUY922 significantly increased the abundance of proteins involved in oxidative phosphorylation and fatty acid metabolism in the PDEs. Consistent with these findings, AUY922-treated prostate cancer cell lines exhibited increased mitochondrial mass and activated fatty acid metabolism processes. We hypothesized that activation of fatty acid oxidation is a potential adaptive response to AUY922 treatment and that cotargeting this process will sensitize prostate cancer cells to HSP90 inhibition. Combination treatment of AUY922 with a clinical inhibitor of fatty acid oxidation, perhexiline, synergistically decreased viability of several prostate cancer cell lines, and had significant efficacy in PDEs. The novel drug combination treatment induced cell-cycle arrest and apoptosis, and attenuated the heat shock response, a known mediator of HSP90 treatment resistance. This combination warrants further preclinical and clinical investigation as a novel strategy to overcome resistance to HSP90 inhibition. Implications: Metabolic pathways induced in tumor cells by therapeutic agents may be critical, but targetable, mediators of treatment resistance.

Purpose The novel Hsp90 inhibitor SNX-2112 showed broad antitumor activity. However, it was still necessary to optimize the therapeutic dosage of SNX-2112 applied on tumors to obtain effective therapy with minimal dose to reduce toxicity. We investigated the role of low-intensity US in promoting antitumorigenic effect of low doses of SNX-2112 on tongue squamous cell carcinoma. Methods Cell viability was measured using CCK-8 assay or staining with Calcein AM/PI. Relative cumulative levels of SNX-2112 in cells were detected using high-performance liquid chromatography. The production of ROS was analyzed using fluorescence microscope and flow cytometer. Cellular apoptosis was detected using flow cytometer. The expression levels of proteins of the ERS-associated apoptosis signaling pathway were detected using Western blotting analysis. The efficacy and biosafety of SNX-2112 were also investigated in a mouse xenograft model. Results Low-intensity US combined with SNX-2112 exhibited significant antitumor effect, increased the absorption of SNX-2112 by cells even with a low dose, enhanced ROS generation and apoptosis. The combination regimen also inhibited the protein expression of Hsp90 and triggered apoptosis through endoplasmic reticulum stress (ERS) by enhancing PERK, CHOP and Bax protein levels, while downregulating the level of Bcl-2. Additionally, N-acetyl-L-cysteine (NAC), ROS scavenger, was able to reverse these results. Low-intensity US combined with SNX-2112 significantly inhibited tumor growth, prolonged survival of mice, decreased proliferation and promoted apoptosis with no visible damage or abnormalities in major organs in the mouse xenograft model with tongue squamous cell carcinoma. Conclusion The antitumor effects of SNX-2112 were enhanced by low-intensity US. The most probable mechanism was that US sonoporation induced more SNX-2112 delivery to the cells and enhanced ROS production, triggering the ERS-associated apoptosis signaling pathway. Therefore, low-intensity US may increase the efficiency of conventional chemotherapy and reduce the dosage of SNX-2112 required and its side effects.

Objective: Breast cancer is one of the most prevalent malignancies and leading causes of females’ mortality worldwide. Because of resistance to various treatment options, new treatments based on molecular targeting has introduced as noticeable strategies in cancer treatment. In this regard, heat shock protein 90 (Hsp90) inhibitors are proposed as effective anticancer drugs. The goal of the study was to utilize a combination of the doxorubicin (DOX) and NVP-AUY 922 on the MCF-7 breast cancer model to investigate the possible cytotoxic mechanisms. Methods: MCF-7 breast cancer cell line was prepared and treated with various concentrations of DOX and NVP-AUY922 in single-drug treatments. We investigated the growth-inhibitory pattern by MTT assay after continuous exposure to NVP-AUY922 and DOX in order to determine dose-response. Then the combinatorial effects were evaluated in concentrations of 0.5 × IC50, 0.2 × IC50, 1 × IC50 and, 2 × IC50 of each drugs. Based on MTT results of double combinations, low effective doses were selected for Real-time PCR [caspase3 and vascular endothelial growth factor (VEGF)] and caspase 3 enzyme activity. Results: A dose-dependent inhibitory effects were presented with increasing the doses of both drugs in single treatments. The upregulation of caspase 3 and downregulation of VEGF mRNA were observed in double combinations of NVP-AUY922 and DOX versus single treatments. Also, in these combinations in low doses of examined drugs (0.5 × IC50, 0.2 × IC50), higher caspase 3 activity were presented in comparison to single treatments (p

Despite several advances in medicine, ischaemic heart disease remains a major cause of morbidity and mortality. The unravelling of molecular mechanisms underlying disease pathophysiology has revealed targets for pharmacological interventions. However, transfer of these pharmcological possibilities to clinical use has been disappointing. Considering the complexity of ischaemic disease at the cellular and molecular levels, an equally multifaceted treatment approach may be envisioned. The pharmacological principle of ‘one target, one key’ may fall short in such contexts, and optimal treatment may involve one or many agents directed against complementary targets. Here, we introduce a ‘multi‐target approach to cardioprotection’ and propose heat shock protein 90 (HSP90) as a target of interest. We report on a member of a distinct class of HSP90 inhibitor possessing pleiotropic activity, which we found to exhibit potent infarct‐sparing effects.

Herein, a series of HSP90 inhibitor-SN38 conjugates through ester and carbamate linkage in the 20-OH and 10-OH positions of SN38 were developed for improving the tumor-specific penetration and accumulation of SN38 via extracellular HSP90 (eHSP90)-mediated endocytosis. Mechanistic analyses confirmed that these novel conjugates could bind to eHSP90 and be selectively internalized into the tumor cells, which led to prolonged tumor regression in multiple models of cancer. Among all studied conjugates, compound 18b showed excellent in vitro activities, including acceptable HSP90α affinity and potent antitumor activity. Moreover, compound 18b exhibited superior antitumor activity and low toxicity in HCT116 and Capan-1 xenograft models. Pharmacokinetic analyses in HCT116 and Capan-1 xenografts further confirmed that compound 18b treatment could lead to effective cleavage and extended SN38 exposure at tumor sites. All these encouraging data indicate that this compound is a promising new candidate for cancer therapy and merits further chemical and biological evaluation.

Patients with advanced-stage cervical cancer have a high rate of morbidity and mortality, predominantly due to the metastasis of cervical cancer cells. Therefore, the development of novel agents to prevent metastasis is required for improved cervical cancer therapeutics. SNX-2112, a potent and selective Hsp90 inhibitor, is an anticancer candidate in clinical trials for the treatment of some solid tumors and lymphomas. However, the effects of SNX-2112 on the migration and invasion of cervical cancer cells are unclear. This study aimed at exploring the effects of SNX-2112 on the migration and invasion of cervical cancer cells and revealing the underlying molecular mechanisms. We found that SNX-2112 significantly decreased the viability, colony formation ability, and migration of HeLa and U14 cells. The invasiveness of HeLa cells and the proteins involved in metastasis were markedly reduced after SNX-2112 treatment. SNX-2112 inactivated the focal adhesion kinase (FAK) and inhibited the expression levels of matrix metallopeptidase (MMP)-9, MMP-2, SLUG, SNAIL, β-catenin, and Vimentin. Furthermore, SNX-2112 inhibited the protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway in HeLa cells by decreasing the phosphorylation of Akt, mTOR, S6, and eukaryotic translation initiation factor 4E-binding protein 1 (4EBP1). It also reduced the expression levels of the endoplasmic reticulum (ER)-localized molecular chaperones, Calnexin and BiP, and unfolded protein response (UPR)-related proteins IRE1α and PERK, suggesting that SNX-2112 can suppress ER stress and thus, inactivate the UPR in HeLa cells. Taken together, these findings indicate that SNX-2112 may efficiently suppress the proliferation, migration, and invasion of cervical cancer cells by inhibiting the Akt/mTOR pathway and could serve as a candidate drug for the treatment of human cervical cancer.

Purpose Currently, the paucity of classical effective pharmacological drugs to treat esophageal squamous cell carcinoma (ESCC) is a major problem. The c-Myc (MYC) protein is a promising target as it is overexpressed in ESCC. MYC is a sensitive client protein of the heat shock protein 90 (HSP90) and, therefore, targeting the HSP90-MYC axis by inhibition of HSP90 is a potential therapeutic strategy for ESCC. Here, we evaluated the clinical application value of the HSP90 inhibitor (Ganetespib, STA-9090) as an anti-cancer agent for MYC-positive ESCC. Materials and methods We first analyzed ESCC tissue microarrays and clinical tissue samples to determine MYC expression. The relationship between MYC and HSP90 was analyzed by co-immunoprecipitation assays and immunofluorescence. In in vitro cell models, cell growth was analyzed using the CCK-8 kit, and MYC protein expression was analyzed by Western blot. The in vivo antitumor activity of STA-9090 was assessed in two xenograft animal models. Results We demonstrated that MYC-overexpressing ESCC cells were highly sensitive to STA-9090 treatment through suppressing ESCC cell proliferation, cell cycle progression and survival. Moreover, STA-9090 treatment decreased MYC expression, reducing the half-life of the MYC protein. We further established two xenograft mouse models using ESCC cells and clinical ESCC samples to validate the effectiveness of STA-9090 in vivo. In both xenograft models, STA-9090 substantially inhibited the growth of MYC-positive ESCC tumors in vivo. In contrast, STA-9090 treatment demonstrated no beneficial effects in mice with low-MYC expressing ESCC tumors. Conclusion In conclusion, our data support that the HSP90 inhibitor, STA-9090, suppresses the expression of the MYC protein and interferes with HSP90-MYC protein-protein interaction. This, in turn, leads to inhibition of ESCC cell proliferation and promotion of apoptosis in ESCC cells in vitro and reduction of ESCC tumors in vivo. We propose, based on our findings, that STA-9090 is a potential novel therapeutic target for MYC-positive ESCC.

SummaryThis study compared the bioavailability of two pimitespib formulations (Formulations A and B), evaluated the food effect on Formulation A, and evaluated the safety and efficacy of multiple pimitespib doses in patients with solid tumors. This clinical, pharmacological multicenter study had two cohorts and periods. A single dose of Formulation A or B was administered in a crossover design to compare the pharmacokinetics in Cohort 1. In Cohort 2, the effects of fed vs fasting conditions were evaluated among those receiving Formulation A. Subsequently, multiple Formulation A doses were administered to all patients for safety and efficacy assessments. In Cohorts 1 and 2, 12 and 16 patients, respectively, were analyzed for pharmacokinetics. Thirty patients were analyzed for safety and efficacy. Maximum concentration (Cmax), area under the curve (AUC)last, and AUCinf geometric mean ratios for Formulations A and B (90% confidence interval [CI]) were 0.8078 (0.6569–0.9933), 0.7973 (0.6672–0.9529), and 0.8094 (0.6697–0.9782), respectively; 90% CIs were not within the bioequivalence range (0.80–1.25). In Cohort 2, mean Cmax, AUClast, and AUCinf were higher in fed vs fasting conditions. No safety concerns emerged with single or multiple administration. Overall response rate, disease control rate, and median progression-free survival were 0%, 33%, and 1.5 months, respectively. Four patients had stable disease ≥ 5 months. Bioequivalence of the two formulations was unconfirmed. Systemic exposure of Formulation A was approximately 20% less than Formulation B. A high-fat/calorie meal increased the relative pharmacokinetics and bioavailability of a single 160-mg dose. Trial Registration: JapicCTI-184191 (Japan Pharmaceutical Information Center) registered on November 5, 2018.

Among human diseases, cancer has been in the frontlines of drug discovery and development. Despite having several decades of research efforts, therapeutic targeting of cancer is still challenging, which is due to the ability of cancer cells to adapt to the tumor microenvironment, exhibiting resistance to therapeutic drugs, and facilitated altered cancer metabolism. The small molecule inhibitors aimed at targeting a selective pathway are becoming void since cancer cells can activate alternate mechanisms. Despite broad acceptance of the Warburg effect, cellular energy metabolism, which determines the cell fate, is often overlooked for cancer treatment. We reported earlier that mitochondrial chaperone, TRAP-1 acts as a switch for activating the alternate cellular metabolism. Hence, we hypothesized that interfering with TRAP-1 inhibition can target the activation of alternative energy metabolism and sensitize tumor cells to existing chemotherapeutic drugs. We developed a nanocarrier where the iron oxide nanoparticles (IONs) were conjugated to Hsp90 inhibitor, geldanamycin (GA), and the mitochondria localization signal (MLS) peptide. We examined its effect against mitochondrial dynamics and metabolic status of human tumor cells. The synthesized nanocarrier exhibited both stability and target-specific activity and did not show nanoparticle-associated cytotoxicity. However, the nanocarrier treated cancer cells exhibited altered mitochondrial morphology and decreased cellular ATP levels suggesting that selective TRAP-1 targeting interferes with the altered energy metabolism. We present a nanoparticle-based TRAP-1 inhibitor to target tumor metabolism.