ABSTRACT
Castration-resistant prostate cancer (CRPC) is characterized by reactivation of androgen receptor (AR) signaling, in part by elevated expression of AR splice variants (ARv) including ARv7, a constitutively active, ligand binding domain (LBD)-deficient variant whose expression has been correlated with therapeutic resistance and poor prognosis. In a screen to identify small-molecule dual inhibitors of both androgen-dependent and androgen-independent AR gene signatures, we identified the chalcone C86. Binding studies using purified proteins and CRPC cell lysates revealed C86 to interact with Hsp40. Pull-down studies using biotinylated-C86 found Hsp40 present in a multiprotein complex with full-length (FL-) AR, ARv7, and Hsp70 in CRPC cells. Treatment of CRPC cells with C86 or the allosteric Hsp70 inhibitor JG98 resulted in rapid protein destabilization of both FL-AR and ARv, including ARv7, concomitant with reduced FL-AR- and ARv7-mediated transcriptional activity. The glucocorticoid receptor, whose elevated expression in a subset of CRPC also leads to androgen-independent AR target gene transcription, was also destabilized by inhibition of Hsp40 or Hsp70. In vivo, Hsp40 or Hsp70 inhibition demonstrated single-agent and combinatorial activity in a 22Rv1 CRPC xenograft model. These data reveal that, in addition to recognized roles of Hsp40 and Hsp70 in FL-AR LBD remodeling, ARv lacking the LBD remain dependent on molecular chaperones for stability and function. Our findings highlight the feasibility and potential benefit of targeting the Hsp40/Hsp70 chaperone axis to treat prostate cancer that has become resistant to standard antiandrogen therapy.Significance: These findings highlight the feasibility of targeting the Hsp40/Hsp70 chaperone axis to treat CRPC that has become resistant to standard antiandrogen therapy. Cancer Res; 78(14); 4022-35. ©2018 AACR.
Subject(s)
Antineoplastic Agents/pharmacology , HSP40 Heat-Shock Proteins/metabolism , HSP70 Heat-Shock Proteins/metabolism , Molecular Chaperones/metabolism , Prostatic Neoplasms, Castration-Resistant/drug therapy , Prostatic Neoplasms, Castration-Resistant/metabolism , Receptors, Androgen/metabolism , A549 Cells , Alternative Splicing/drug effects , Androgen Antagonists/pharmacology , Androgens/metabolism , Animals , COS Cells , Cell Line , Cell Line, Tumor , Chlorocebus aethiops , HEK293 Cells , Humans , Male , Mice, Nude , RNA Splicing/drug effects , Signal Transduction/drug effects , Transcription, Genetic/drug effectsABSTRACT
Prostate cancer (PC) is the fifth leading cause of cancer death in men, and the androgen receptor (AR) represents the primary target for PC treatment, even though the disease frequently progresses toward androgen-independent forms. Most of the commercially available nonsteroidal antiandrogens show a common scaffold consisting of two aromatic rings connected by a linear or a cyclic spacer. By taking advantage of a facile, one-pot click chemistry reaction, we report herein the preparation of a small library of novel 1,4-substituted triazoles with AR antagonistic activity. Biological and theoretical evaluation demonstrated that the introduction of the triazole core in the scaffold of nonsteroidal antiandrogens allowed the development of small molecules with improved overall AR-antagonist activity. In fact, compound 14d displayed promising in vitro antitumor activity toward three different prostate cancer cell lines and was able to induce 60% tumor growth inhibition of the CW22Rv1 in vivo xenograft model. These results represent a step toward the development of novel and improved AR antagonists.
Subject(s)
Nonsteroidal Anti-Androgens/chemistry , Nonsteroidal Anti-Androgens/therapeutic use , Prostate/drug effects , Prostatic Neoplasms/drug therapy , Triazoles/chemistry , Triazoles/therapeutic use , Animals , Cell Line, Tumor , Drug Discovery , Gene Expression Regulation, Neoplastic/drug effects , Humans , Male , Mice, Nude , Models, Molecular , Nonsteroidal Anti-Androgens/pharmacology , Prostate/metabolism , Prostate/pathology , Prostate-Specific Antigen/genetics , Prostatic Neoplasms/genetics , Prostatic Neoplasms/pathology , RNA, Messenger/genetics , Triazoles/pharmacologyABSTRACT
SUMO-modification of nuclear proteins has profound effects on gene expression. However, non-toxic chemical tools that modulate sumoylation in cells are lacking. Here, to identify small molecule sumoylation inhibitors we developed a cell-based screen that focused on the well-sumoylated substrate, human Liver Receptor Homolog-1 (hLRH-1, NR5A2). Our primary gene-expression screen assayed two SUMO-sensitive transcripts, APOC3 and MUC1, that are upregulated by SUMO-less hLRH-1 or by siUBC9 knockdown, respectively. A polyphenol, tannic acid (TA) emerged as a potent sumoylation inhibitor in vitro (IC50 = 12.8 µM) and in cells. TA also increased hLRH-1 occupancy on SUMO-sensitive transcripts. Most significantly, when tested in humanized mouse primary hepatocytes, TA inhibits hLRH-1 sumoylation and induces SUMO-sensitive genes, thereby recapitulating the effects of expressing SUMO-less hLRH-1 in mouse liver. Our findings underscore the benefits of phenotypic screening for targeting post-translational modifications, and illustrate the potential utility of TA for probing the cellular consequences of sumoylation.
Subject(s)
Enzyme Inhibitors/isolation & purification , Enzyme Inhibitors/metabolism , Hepatocytes/drug effects , Receptors, Cytoplasmic and Nuclear/metabolism , Sumoylation/drug effects , Tannins/isolation & purification , Tannins/metabolism , Animals , Cells, Cultured , Drug Evaluation, Preclinical/methods , Gene Expression Profiling , Hepatocytes/enzymology , Humans , Inhibitory Concentration 50 , Mice , Mice, SCIDABSTRACT
Protein sumoylation is a dynamic posttranslational modification that regulates a diverse subset of the proteome. The mechanism by which sumoylation enzymes recognize their cognate substrates is unclear, and the consequences of sumoylation remain difficult to predict. While small molecule probes of the sumoylation process could be valuable for understanding SUMO biology, few small molecules that modulate this process exist. Here, we report the synthesis and evaluation of over 600 oxime-containing peptide sumoylation substrates. Our work demonstrates that higher modification efficiency can be achieved with non-natural side chains that deviate substantially from the consensus site requirement of a hydrophobic substituent. Furthermore, docking studies suggest that these improved substrates mimic binding interactions that are used by other endogenous protein sequences through tertiary interactions. The development of these high efficiency substrates provides key mechanistic insights toward specific recognition of low molecular weight species in the sumoylation pathway.
Subject(s)
Consensus Sequence , Small Ubiquitin-Related Modifier Proteins/chemistry , Sumoylation , Substrate SpecificityABSTRACT
Protein sumoylation is a dynamic posttranslational modification involved in diverse biological processes during cellular homeostasis and development. Recently sumoylation has been shown to play a critical role in cancer, although to date there are few small molecule probes available to inhibit enzymes involved in the SUMO conjugation process. As part of a program to identify and study inhibitors of sumoylation we recently reported the discovery that 2',3',4'-trihydroxyflavone (2-D08) is a cell permeable, mechanistically unique inhibitor of protein sumoylation. The work reported herein describes an efficient synthesis of 2-D08 as well as a structurally related but inactive isomer. We also report an unanticipated Wessely-Moser rearrangement that occurs under vigorous methyl ether deprotection conditions. This rearrangement likely gave rise to 2-D08 during a deprotection step, resulting in 2-D08 appearing as a contaminant in a screening well from a commercial supplier.
Subject(s)
Flavones/pharmacology , Small Ubiquitin-Related Modifier Proteins/antagonists & inhibitors , Flavones/chemical synthesis , Flavones/chemistry , Small Ubiquitin-Related Modifier Proteins/metabolism , Sumoylation/drug effectsABSTRACT
A library of 3-hydroxy-2,3-dihydropyridones was synthesized, and their activities as antiandrogens were tested in the human prostate cancer cell line LNCaP. Structure-activity relationship (SAR) studies resulted in the identification of a potent compound whose activity is comparable to that of MDV3100. Homology modeling and molecular mechanics were used to build a structural model of the androgen receptor-ligand binding domain and to investigate the structural basis of the antagonism. The model is qualitatively consistent with the observed SAR. Moreover, the enrichment plot shows that screening with the model performs significantly better than random screening. Therefore, the model probably represents a realistic conformation of the antagonist form and can be utilized for structure-based design of novel antiandrogens.
Subject(s)
Pyridones/chemical synthesis , Pyridones/pharmacology , Receptors, Androgen/metabolism , Cell Line, Tumor , Crystallography, X-Ray , Dose-Response Relationship, Drug , Humans , Models, Molecular , Molecular Structure , Pyridones/chemistry , Receptors, Androgen/chemistry , Stereoisomerism , Structure-Activity RelationshipABSTRACT
The dynamic, posttranslational modification of proteins with a small ubiquitin-like modifier (SUMO) tag has been recognized as an important cellular regulatory mechanism relevant to a number of cancers as well as normal embryonic development. As part of a program aimed toward the identification of inhibitors of SUMO-conjugating enzymes, we developed a microfluidic electrophoretic mobility shift assay to monitor sumoylation events in real time. We disclose herein the use of this assay to identify a cell-permeable compound capable of blocking the transfer of SUMO-1 from the E2 enzyme Ubc9 to the substrate. We screened a small collection of compounds and identified an oxygenated flavonoid derivative that inhibits sumoylation in vitro. Next, we carried out an in-depth mechanistic analysis that ruled out many common false-positive mechanisms such as aggregation or alkylation. Furthermore, we report that this flavonoid inhibits a single step in the sumoylation cascade: the transfer of SUMO from the E2 enzyme (Ubc9) thioester conjugate to the substrate. In addition to having a unique mechanism of action, this inhibitor has a discrete structure-activity relationship uncharacteristic of a promiscuous inhibitor. Cell-based studies showed that the flavonoid inhibits the sumoylation of topoisomerase-I in response to camptothecin treatment in two different breast cancer cell lines, while isomeric analogs are inactive. Importantly, this compound blocks sumoylation while not affecting ubiquitylation in cells. This work identifies a point of entry for pharmacologic inhibition of the sumoylation cascade and may serve as the basis for continued study of additional pharmacophores that modulate SUMO-conjugating enzymes such as Ubc9.
Subject(s)
Electrophoretic Mobility Shift Assay , SUMO-1 Protein/metabolism , Small Molecule Libraries/chemistry , Amino Acid Sequence , Cell Line, Tumor , Humans , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics , SUMO-1 Protein/antagonists & inhibitors , SUMO-1 Protein/genetics , Small Molecule Libraries/pharmacology , Structure-Activity Relationship , Sumoylation/drug effectsABSTRACT
Androgen receptor activity drives incurable castrate-resistant prostate cancer. All approved antiandrogens inhibit androgen receptor-driven transcription, and in addition the second-generation antiandrogen MDV3100 inhibits ligand-activated androgen receptor nuclear translocation, via an unknown mechanism. Here, we report methoxychalcones that lock the heat shock protein 90-androgen receptor complex in the cytoplasm in an androgen-non-responsive state, thus demonstrating a novel chemical scaffold for antiandrogen development and a unique mechanism of antiandrogen activity.
Subject(s)
Androgen Antagonists/chemistry , Androgen Antagonists/pharmacology , Chalcones/chemistry , Chalcones/pharmacology , Receptors, Androgen/metabolism , Cell Line, Tumor , Gene Expression Regulation, Neoplastic/drug effects , HSP90 Heat-Shock Proteins/metabolism , Humans , Male , Prostatic Neoplasms/drug therapy , Prostatic Neoplasms/genetics , Prostatic Neoplasms/metabolism , Protein Transport/drug effects , Receptors, Androgen/analysis , Receptors, Androgen/geneticsABSTRACT
Inactivation of the TCA cycle enzyme, fumarate hydratase (FH), drives a metabolic shift to aerobic glycolysis in FH-deficient kidney tumors and cell lines from patients with hereditary leiomyomatosis renal cell cancer (HLRCC), resulting in decreased levels of AMP-activated kinase (AMPK) and p53 tumor suppressor, and activation of the anabolic factors, acetyl-CoA carboxylase and ribosomal protein S6. Reduced AMPK levels lead to diminished expression of the DMT1 iron transporter, and the resulting cytosolic iron deficiency activates the iron regulatory proteins, IRP1 and IRP2, and increases expression of the hypoxia inducible factor HIF-1α, but not HIF-2α. Silencing of HIF-1α or activation of AMPK diminishes invasive activities, indicating that alterations of HIF-1α and AMPK contribute to the oncogenic growth of FH-deficient cells.
Subject(s)
AMP-Activated Protein Kinases/metabolism , Fumarate Hydratase/deficiency , Iron Deficiencies , Kidney Neoplasms/metabolism , Leiomyomatosis/congenital , Acetyl Coenzyme A/biosynthesis , Acetyl-CoA Carboxylase/biosynthesis , Acetyl-CoA Carboxylase/metabolism , Animals , Basic Helix-Loop-Helix Transcription Factors/biosynthesis , Cation Transport Proteins/biosynthesis , Cell Line, Tumor , Fumarate Hydratase/metabolism , Glycolysis/drug effects , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/biosynthesis , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Iron Regulatory Protein 1/biosynthesis , Iron Regulatory Protein 1/metabolism , Iron Regulatory Protein 2/biosynthesis , Iron Regulatory Protein 2/metabolism , Kidney Neoplasms/enzymology , Kidney Neoplasms/pathology , Leiomyomatosis/metabolism , Leiomyomatosis/pathology , Mice , NADP/biosynthesis , Neoplastic Syndromes, Hereditary , Ribose/biosynthesis , Ribosomal Protein S6/biosynthesis , Ribosomal Protein S6/metabolism , Skin Neoplasms , Thenoyltrifluoroacetone/pharmacology , Tumor Suppressor Protein p53/biosynthesis , Uterine NeoplasmsABSTRACT
PURPOSE: To determine the maximum tolerated dose (MTD), toxicities, and pharmacokinetic/pharmacodynamic profile of the Hsp90 inhibitor PF-04929113 (SNX-5422) in patients with advanced solid tumors and lymphomas. METHODS: This was a single-institution, phase I, dose-escalation study of PF-04929113 administered twice weekly. Endpoints included determination of dose-limiting toxicities (DLT), MTD, the safety profile of PF-04929113, pharmacodynamic assessment of PF-04929113 on Hsp70 induction, pharmacokinetic analysis of PF-04928473 (SNX-2112) and its prodrug PF-04929113, and assessment of response. RESULTS: Thirty-three patients with advanced malignancies were treated. Dose escalation was continued up to 177 mg/m(2) administered orally twice a week. One DLT (nonseptic arthritis) was noted. No grade 4 drug-related adverse events were seen; grade 3 adverse events included diarrhea (9%), nonseptic arthritis (3%), aspartate aminotransferase elevation (3%), and thrombocytopenia (3%). No objective responses were seen in 32 evaluable patients. Fifteen patients (47%) had stable disease; 17 patients (53%) had progressive disease. Pharmacokinetic data revealed rapid absorption, hepatic, and extrahepatic clearance, extensive tissue binding, and almost linear pharmacokinetics of the active drug PF-04928473. Pharmacodynamic studies confirmed inhibition of Hsp90 and a linear correlation between pharmacokinetic parameters and Hsp70 induction. CONCLUSIONS: PF-04929113 administered orally twice a week is well tolerated and inhibits its intended target Hsp90. No objective responses were seen, but long-lasting stabilizations were obtained. Although no clinically significant drug-related ocular toxicity was seen in this study, the development of PF-04929113 has been discontinued because of ocular toxicity seen in animal models and in a separate phase I study.
Subject(s)
Benzamides/administration & dosage , Benzamides/adverse effects , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Indazoles/administration & dosage , Indazoles/adverse effects , Lymphoma/drug therapy , Neoplasms/drug therapy , Adult , Aged , Benzamides/pharmacokinetics , Dose-Response Relationship, Drug , Drug Administration Schedule , Female , Glycine , HSP90 Heat-Shock Proteins/blood , Humans , Indazoles/pharmacokinetics , Lymphoma/blood , Lymphoma/metabolism , Male , Middle Aged , Neoplasms/blood , Neoplasms/metabolismABSTRACT
Drugs that target novel surfaces on the androgen receptor (AR) and/or novel AR regulatory mechanisms are promising alternatives for the treatment of castrate-resistant prostate cancer. The 52 kDa FK506 binding protein (FKBP52) is an important positive regulator of AR in cellular and whole animal models and represents an attractive target for the treatment of prostate cancer. We used a modified receptor-mediated reporter assay in yeast to screen a diversified natural compound library for inhibitors of FKBP52-enhanced AR function. The lead compound, termed MJC13, inhibits AR function by preventing hormone-dependent dissociation of the Hsp90-FKBP52-AR complex, which results in less hormone-bound receptor in the nucleus. Assays in early and late stage human prostate cancer cells demonstrated that MJC13 inhibits AR-dependent gene expression and androgen-stimulated prostate cancer cell proliferation.
Subject(s)
Gene Expression Regulation/drug effects , HSP90 Heat-Shock Proteins/metabolism , Models, Molecular , Multiprotein Complexes/metabolism , Prostatic Neoplasms/metabolism , Receptors, Androgen/metabolism , Tacrolimus Binding Proteins/antagonists & inhibitors , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Discovery , Enzyme-Linked Immunosorbent Assay , Fluorescence , Humans , Immunoblotting , Immunoprecipitation , Male , Mice , Molecular Dynamics Simulation , Molecular Structure , Receptors, Androgen/chemistry , Tacrolimus Binding Proteins/metabolism , Yeasts , beta-GalactosidaseABSTRACT
The molecular chaperone Heat Shock Protein 90 (Hsp90) is essential for the function of various oncoproteins that are vital components of multiple signaling networks regulating cancer cell proliferation, survival, and metastasis. Hsp90 chaperone function is coupled to its ATPase activity, which can be inhibited by natural products such as the ansamycin geldanamycin (GA) and the resorcinol radicicol (RD). These compounds have served as templates for development of numerous natural product Hsp90 inhibitors. More recently, second generation, fully synthetic Hsp90 inhibitors, based on a variety of chemical scaffolds, have also been synthesized. Together, 18 natural product and synthetic Hsp90 inhibitors have entered clinical trial in cancer patients. To successfully develop Hsp90 inhibitors for oncology indications it is important to understand the factors that influence the susceptibility of Hsp90 to these drugs in vivo. We recently reported that Casein Kinase 2 phosphorylates a conserved threonine residue (T22) in helix-1 of the yeast Hsp90 N-domain both in vitro and in vivo. Phosphorylation of this residue reduces ATPase activity and affects Hsp90 chaperone function. Here, we present additional data demonstrating that ATP binding but not N-domain dimerization is a prerequisite for T22 phosphorylation. We also provide evidence that T22 is an important determinant of Hsp90 inhibitor sensitivity in yeast and we show that T22 phosphorylation status contributes to drug sensitivity in vivo.
Subject(s)
Casein Kinase II/metabolism , Fungal Proteins/metabolism , Lactams, Macrocyclic/therapeutic use , Neoplasms/drug therapy , Resorcinols/therapeutic use , Clinical Trials as Topic , Drug Design , Drug Resistance , HSP90 Heat-Shock Proteins/metabolism , Humans , Molecular Chaperones/metabolism , Molecular Targeted Therapy , Phosphorylation , Threonine/metabolism , YeastsABSTRACT
PURPOSE: Inhibition of epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) pathways may result in synergistic antitumour activity. We designed a phase I study to evaluate the combination of vandetanib, an investigational agent with activity against EGF receptor and VEGF receptor 2, and bevacizumab, a monoclonal antibody against VEGF. EXPERIMENTAL DESIGN: Patients with advanced solid tumours and lymphomas were enrolled. Objectives were to determine the safety and maximum tolerated dose of the combination, characterise pharmacokinetics, measure angiogenic marker changes in blood, and assess tumour blood flow using dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). Vandetanib was given orally once daily and bevacizumab intravenously once in every 3 weeks in 21-day cycles utilising a standard dose-escalation design. RESULTS: Fifteen patients were enrolled, and a total of 94 cycles of therapy were administered. No protocol-defined dose-limiting toxicities were observed; due to toxicities associated with chronic dosing, hypertension, proteinuria, diarrhoea and anorexia, dose escalation was stopped at the second dose level. We observed one partial response and one minor response; 9 patients experienced stable disease. There were significant changes in plasma VEGF and placental-derived growth factor levels, and decreases in K(trans) and k(ep) were observed by DCE-MRI. CONCLUSION: In this trial, we safely combined two targeted agents that cause dual blockade of the VEGF pathway, demonstrated preliminary evidence of clinical activity, and conducted correlative studies demonstrating anti-angiogenic effect. The recommended phase II dose was established as vandetanib 200 mg daily and bevacizumab 7.5 mg/kg every 3 weeks.
Subject(s)
Antibodies, Monoclonal/therapeutic use , Epidermal Growth Factor/metabolism , Lymphoma/drug therapy , Neoplasms/drug therapy , Piperidines/therapeutic use , Quinazolines/therapeutic use , Vascular Endothelial Growth Factor A/metabolism , Adult , Aged , Antibodies, Monoclonal, Humanized , Antineoplastic Agents/pharmacology , Bevacizumab , Biomarkers, Tumor , Contrast Media/pharmacology , Epidermal Growth Factor/antagonists & inhibitors , Female , Humans , Lymphoma/metabolism , Magnetic Resonance Imaging/methods , Male , Middle Aged , Neoplasms/metabolism , Piperidines/pharmacology , Quinazolines/pharmacology , Signal Transduction , Time Factors , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor Receptor-2/metabolismABSTRACT
PURPOSE We previously demonstrated that thalidomide appears to add to the activity of docetaxel in metastatic castration-resistant prostate cancer (CRPC). Phase II studies combining docetaxel with bevacizumab have had substantial antitumor activity. We hypothesized that the combination of docetaxel plus these antiangiogenic drugs with different targets would have substantial clinical activity. To explore safety and efficacy, this was tested in mice and in human patients. PATIENTS AND METHODS Preclinical efficacy of the combination therapy was evaluated in PC3 xenograft mice. Sixty patients with progressive metastatic CRPC received intravenous docetaxel and bevacizumab plus oral thalidomide and prednisone. The primary end point was a prostate-specific antigen (PSA) decline of > or = 50%. Secondary end points included time to progression, overall survival, and safety. Results In the mouse model, combination therapy of docetaxel, bevacizumab, and thalidomide inhibited tumor growth most effectively. In the clinical trial, 90% of patients receiving the combination therapy had PSA declines of > or = 50%, and 88% achieved a PSA decline of > or = 30% within the first 3 months of treatment. The median time to progression was 18.3 months, and the median overall survival was 28.2 months in this group with a Halabi-predicted survival of 14 months. While toxicities were manageable, all patients developed grade 3/4 neutropenia. CONCLUSION The addition of bevacizumab and thalidomide to docetaxel is a highly active combination with manageable toxicities. The estimated median survival is encouraging, given the generally poor prognosis of this patient population. These results suggest that definitive clinical trials combining antiangiogenic agent combinations with docetaxel are warranted to improve treatment outcomes for patients with metastatic CRPC.
Subject(s)
Adenocarcinoma/drug therapy , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Orchiectomy , Prostatic Neoplasms/drug therapy , Adenocarcinoma/immunology , Adenocarcinoma/secondary , Adenocarcinoma/surgery , Administration, Oral , Adult , Aged , Angiogenesis Inhibitors/administration & dosage , Animals , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal, Humanized , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/pharmacokinetics , Bevacizumab , Cell Line, Tumor , Disease-Free Survival , Docetaxel , Down-Regulation , Humans , Infusions, Intravenous , Kaplan-Meier Estimate , Male , Mice , Mice, SCID , Middle Aged , Prednisone/administration & dosage , Prostate-Specific Antigen/blood , Prostatic Neoplasms/immunology , Prostatic Neoplasms/pathology , Prostatic Neoplasms/surgery , Taxoids/administration & dosage , Thalidomide/administration & dosage , Time Factors , Treatment Outcome , Tumor Burden/drug effects , Xenograft Model Antitumor AssaysABSTRACT
To identify pathways controlling prostate cancer metastasis we performed differential display analysis of the human prostate carcinoma cell line PC-3 and its highly metastatic derivative PC-3M. This revealed that a 78-kDa interferon-inducible GTPase, MxA, was expressed in PC-3 but not in PC-3M cells. The gene encoding MxA, MX1, is located in the region of chromosome 21 deleted as a consequence of fusion of TMPRSS2 and ERG, which has been associated with aggressive, invasive prostate cancer. Stable exogenous MxA expression inhibited in vitro motility and invasiveness of PC-3M cells. In vivo exogenous MxA expression decreased the number of hepatic metastases following intrasplenic injection. Exogenous MxA also reduced motility and invasiveness of highly metastatic LOX melanoma cells. A mutation in MxA that inactivated its GTPase reversed inhibition of motility and invasion in both tumor cell lines. Co-immunoprecipitation studies demonstrated that MxA associated with tubulin, but the GTPase-inactivating mutation blocked this association. Because MxA is a highly inducible gene, an MxA-targeted drug discovery screen was initiated by placing the MxA promoter upstream of a luciferase reporter. Examination of the NCI diversity set of small molecules revealed three hits that activated the promoter. In PC-3M cells, these drugs induced MxA protein and inhibited motility. These data demonstrate that MxA inhibits tumor cell motility and invasion, and that MxA expression can be induced by small molecules, potentially offering a new approach to the prevention and treatment of metastasis.
Subject(s)
Cell Movement/drug effects , GTP-Binding Proteins/metabolism , Interferon-alpha/pharmacology , Neoplasms/pathology , Amino Acid Substitution/genetics , Animals , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Screening Assays, Antitumor , GTP-Binding Proteins/chemistry , GTP-Binding Proteins/genetics , Gene Expression Regulation, Neoplastic/drug effects , Genome, Human/genetics , Humans , Liver Neoplasms/secondary , Mice , Microtubules/drug effects , Microtubules/enzymology , Mutant Proteins/metabolism , Mutation/genetics , Myxovirus Resistance Proteins , Neoplasm Invasiveness , Protein Binding/drug effects , Small Molecule Libraries/analysis , Time Factors , Tubulin/metabolismABSTRACT
PURPOSE OF REVIEW: The purpose of this review is to provide an overview of recent advances in the development of histone deacetylase inhibitors (HDACi) for the treatment of cancer. RECENT FINDINGS: Recently, there has been a dramatic expansion of HDACi clinical investigation. There are now 11 HDACi in clinical trial, including inhibitors with a broad spectrum of HDAC isoform inhibitory activity as well as drugs with isoform selectivity. Over 70 combination therapy trials are in progress. Major areas of progress covered include the entry of new HDAC inhibitors into clinical development, recent progress in understanding of molecular mechanisms of HDACi anticancer activity, and a preclinical and clinical update on HDACi in combination. SUMMARY: In the period under review there have been advances in understanding of HDACi mechanisms of action, identification of rational combinations that address increased efficacy and overcoming resistance, and greatly expanded clinical development of pan-HDAC-inhibitory and isoform-selective inhibitors in monotherapy and combination therapy protocols.
Subject(s)
Antineoplastic Agents/therapeutic use , Histone Deacetylase Inhibitors , Neoplasms/drug therapy , Neoplasms/pathology , Apoptosis , Chemistry, Pharmaceutical/methods , Chemistry, Pharmaceutical/trends , Clinical Trials as Topic , Drug Design , Enzyme Inhibitors/therapeutic use , Histones/metabolism , Humans , Immune System , Medical Oncology/methods , Medical Oncology/trends , Models, Chemical , Neoplastic Stem Cells , Protein IsoformsABSTRACT
Infrared spectroscopy identifies molecules by detection of vibrational patterns characteristic of molecular bonds. We apply this approach to measure protein acetylation after treatment with histone deacetylase inhibitors. The anticancer activity of histone deacetylase inhibitors (HDACi) is ascribed to the hyperacetylation of both core nucleosomal histones and nonhistone proteins critical to the maintenance of the malignant phenotype (Marks, P. A.; Richon, V. M.; Breslow, R.; Rifkind, R. A. Curr. Opin. Oncol. 2001, 13, 477-483; Mai, A.; Massa, S.; Rotili, D.; Cerbara, I.; Valente, S.; Pezzi, R.; Simeoni, S.; Ragno, R. Med. Res. Rev. 2005, 25, 261-309). After incubation of the peripheral blood mononuclear cells (PBMCs) in vitro with the HDACi SNDX-275, a benzamide drug derivative, vibrational spectral changes in the methyl and methylene stretching mode regions, which reflect concentration-dependent increases in protein acetylation, were detected and quantified. We applied these metrics, based upon spectral differences, to peripheral blood mononuclear cells from patients treated in vivo with this agent. The data demonstrate a new approach to a sensitive assessment of global molecular modifications that is independent of antibodies, requires minimal cell processing, and is easily adapted to high-throughput screening.
Subject(s)
Benzamides/pharmacology , Diagnostic Imaging/methods , Enzyme Inhibitors/pharmacology , Histone Deacetylase Inhibitors , Histone Deacetylases/chemistry , Leukocytes, Mononuclear/cytology , Pyridines/pharmacology , Spectroscopy, Fourier Transform Infrared/methods , Acetylation , Cells, Cultured , Histone Deacetylases/metabolism , Humans , VibrationSubject(s)
Acetylene/analogs & derivatives , Benzene Derivatives/chemistry , Molybdenum/chemistry , Nanostructures/chemistry , Oxides/chemistry , Polymers/chemistry , Rotaxanes/chemistry , Acetylene/chemical synthesis , Acetylene/chemistry , Benzene Derivatives/chemical synthesis , Models, Molecular , Molecular Structure , Particle Size , Surface PropertiesABSTRACT
A new series of hematoporphyrin-platinum(II) conjugates was prepared by platination of the glutamate ligand tethered to hydrophilic hematoporphyrin derivatives, in which different numbers of ethylene oxide unit were introduced to modulate the hydrophobic/hydrophilic balance of the conjugates. The antitumor activity of the hematoporphyrin-platinum(II) conjugates was assayed in vitro and in vivo against the leukemia L1210 cell line. Among the complexes, compound 11 exhibited not only higher in vivo activity (T/C% = 192) than cisplatin (T/C% = 184) and carboplatin (T/C% = 168), but also elevated tumor-localizing effect (tumor/muscle ratio > 3).