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Metabolic rewiring promotes cancer cell adaptation to a hostile microenvironment, representing a hallmark of cancer. This process involves mitochondrial function and is mechanistically linked to the balance between mitochondrial biogenesis (MB) and mitophagy. The molecular chaperone TRAP1 is overexpressed in 60-70% of human colorectal cancers (CRC) and its over-expression correlates with poor clinical outcome, being associated with many cancer cell functions (i.e. adaptation to stress, protection from apoptosis and drug resistance, protein synthesis quality control, metabolic rewiring from glycolysis to mitochondrial respiration and vice versa). Here, the potential new role of TRAP1 in regulating mitochondrial dynamics was investigated in CRC cell lines and human CRCs. Our results revealed an inverse correlation between TRAP1 and mitochondrial-encoded respiratory chain proteins both at transcriptional and translational levels. Furthermore, TRAP1 silencing is associated with increased mitochondrial mass and mitochondrial DNA copy number (mtDNA-CN) as well as enhanced MB through PGC-1α/TFAM signalling pathway, promoting the formation of new functioning mitochondria and, likely, underlying the metabolic shift towards oxidative phosphorylation. These results suggest an involvement of TRAP1 in regulating MB process in human CRC cells. KEY MESSAGES: TRAP1 inversely correlates with protein-coding mitochondrial gene expression in CRC cells and tumours. TRAP1 silencing correlates with increased mitochondrial mass and mtDNA copy number in CRC cells. TRAP1 silencing favours mitochondrial biogenesis in CRC cells., (© 2024. The Author(s).)

Metabolic rewiring is a mechanism of adaptation to unfavorable environmental conditions and tumor progression. TRAP1 is an HSP90 molecular chaperone upregulated in human colorectal carcinomas (CRCs) and responsible for downregulation of oxidative phosphorylation (OXPHOS) and adaptation to metabolic stress. The mechanism by which TRAP1 regulates glycolytic metabolism and the relevance of this regulation in resistance to EGFR inhibitors were investigated in patient-derived CRC spheres, human CRC cells, samples, and patients. A linear correlation was observed between TRAP1 levels and 18 F-fluoro-2-deoxy-glucose ( 18 F-FDG) uptake upon PET scan or GLUT1 expression in human CRCs. Consistently, TRAP1 enhances GLUT1 expression, glucose uptake, and lactate production and downregulates OXPHOS in CRC patient-derived spheroids and cell lines. Mechanistically, TRAP1 maximizes lactate production to balance low OXPHOS through the regulation of the glycolytic enzyme phosphofructokinase-1 (PFK1); this depends on the interaction between TRAP1 and PFK1, which favors PFK1 glycolytic activity and prevents its ubiquitination/degradation. By contrast, TRAP1/PFK1 interaction is lost in conditions of enhanced OXPHOS, which results in loss of TRAP1 regulation of PFK1 activity and lactate production. Notably, TRAP1 regulation of glycolysis is involved in resistance of RAS-wild-type CRCs to EGFR monoclonals. Indeed, either TRAP1 upregulation or high glycolytic metabolism impairs cetuximab activity in vitro, whereas TRAP1 targeting and/or inhibition of glycolytic pathway enhances cell response to cetuximab. Finally, a linear correlation between 18 F-FDG PET uptake and poor response to cetuximab in first-line therapy in human metastatic CRCs was observed. These results suggest that TRAP1 is a key determinant of CRC metabolic rewiring and favors resistance to EGFR inhibitors through regulation of glycolytic metabolism., (© 2020 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.)

Wnt/β-Catenin signaling is involved in embryonic development, regeneration, and cellular differentiation and is responsible for cancer stemness maintenance. The HSP90 molecular chaperone TRAP1 is upregulated in 60-70% of human colorectal carcinomas (CRCs) and favors stem cells maintenance, modulating the Wnt/β-Catenin pathway and preventing β-Catenin phosphorylation/degradation. The role of TRAP1 in the regulation of Wnt/β-Catenin signaling was further investigated in human CRC cell lines, patient-derived spheroids, and CRC specimens. TRAP1 relevance in the activation of Wnt/β-Catenin signaling was highlighted by a TCF/LEF Cignal Reporter Assay in Wnt-off HEK293T and CRC HCT116 cell lines. Of note, this regulation occurs through the modulation of Wnt ligand receptors LRP5 and LRP6 that are both downregulated in TRAP1-silenced cell lines. However, while LRP5 mRNA is significantly downregulated upon TRAP1 silencing, LRP6 mRNA is unchanged, suggesting independent mechanisms of regulation by TRAP1. Indeed, LRP5 is regulated upon promoter methylation in CRC cell lines and human CRCs, whereas LRP6 is controlled at post-translational level by protein ubiquitination/degradation. Consistently, human CRCs with high TRAP1 expression are characterized by the co-upregulation of active β-Catenin, LRP5 and LRP6. Altogether, these data suggest that Wnt/β-Catenin signaling is modulated at multiple levels by TRAP1.

Tumor Necrosis Factor Receptor-Associated Protein 1 (TRAP1) is a heat shock protein 90 (HSP90) molecular chaperone overexpressed in 60-70% human colorectal carcinomas (CRCs) and the co-upregulation of TRAP1 and associated 6-related proteins identifies metastatic CRCs with poor prognosis. Since the molecular mechanisms responsible for TRAP1 regulation are still unknown, the significance of TRAP1 gene copy number (CN) and the role of post-transductional protein modifications were addressed. TRAP1 gene aneuploidy accounted for 34.5% of cases in a cohort of 58 human CRCs and TRAP1 CN correlated with its mRNA and protein expression, suggesting that transcriptional mechanisms are responsible for TRAP1 upregulation. Furthermore, the analysis of the National Cancer Institute's Clinical Proteomic Tumor Analysis Consortium/The Cancer Genome Atlas (CPTAC/TCGA) CRC database showed that TRAP1 polysomy significantly correlates with lymph node involvement. However, a subgroup of tumors showed TRAP1 protein levels independent from its CN. Of note, a direct correlation was observed between TRAP1 protein levels and the expression of S-nitrosoglutathione reductase (GSNOR), a denitrosylase involved in the regulation of protein S-nitrosylation. Furthermore, CRC cell lines exposed to hypoxia or dichloroacetate treatment showed the downregulation of TRAP1 upon GSNOR silencing and this resulted in increased TRAP1 mono/polyubiquitination. These data suggest that transcriptional and post-transductional mechanisms account for TRAP1 expression in human CRCs and GSNOR protects TRAP1 from S-nitrosylation and consequent proteasome degradation mostly in conditions of stress.

Heat shock protein 90 (HSP90) molecular chaperones are a family of ubiquitous proteins participating in several cellular functions through the regulation of folding and/or assembly of large multiprotein complexes and client proteins. Thus, HSP90s chaperones are, directly or indirectly, master regulators of a variety of cellular processes, such as adaptation to stress, cell proliferation, motility, angiogenesis, and signal transduction. In recent years, it has been proposed that HSP90s play a crucial role in carcinogenesis as regulators of genotype-to-phenotype interplay. Indeed, HSP90 chaperones control metabolic rewiring, a hallmark of cancer cells, and influence the transcription of several of the key-genes responsible for tumorigenesis and cancer progression, through either direct binding to chromatin or through the quality control of transcription factors and epigenetic effectors. In this review, we will revise evidence suggesting how this interplay between epigenetics and metabolism may affect oncogenesis. We will examine the effect of metabolic rewiring on the accumulation of specific metabolites, and the changes in the availability of epigenetic co-factors and how this process can be controlled by HSP90 molecular chaperones. Understanding deeply the relationship between epigenetic and metabolism could disclose novel therapeutic scenarios that may lead to improvements in cancer treatment., Competing Interests: The authors declare no conflict of interest.

TRAP1 (Hsp75) is the mitochondrial paralog of the Hsp90 molecular chaperone family. Due to structural similarity among Hsp90 chaperones, a potential strategy to induce apoptosis through mitochondrial TRAP1 ATPase inhibition has been envisaged and a series of compounds has been developed by binding the simple pharmacophoric core of known Hsp90 inhibitors with various appendages bearing a permanent cationic head, or a basic group highly ionizable at physiologic pH. Cationic appendages were selected as vehicles to deliver drugs to mitochondria. Indeed, masses of new derivatives were evidenced to accumulate in the mitochondrial fraction from colon carcinoma cells and a compound in the series, with a guanidine appendage, demonstrated good activity in inhibiting recombinant TRAP1 ATPase and cell growth and in inducing apoptotic cell death in colon carcinoma cells., (Copyright © 2018. Published by Elsevier Ltd.)

Regulation of tumour cell proliferation by molecular chaperones is still a complex issue. Here, the role of the HSP90 molecular chaperone TRAP1 in cell cycle regulation was investigated in a wide range of human breast, colorectal, and lung carcinoma cell lines, and tumour specimens. TRAP1 modulates the expression and/or the ubiquitination of key cell cycle regulators through a dual mechanism: (i) transcriptional regulation of CDK1, CYCLIN B1, and MAD2, as suggested by gene expression profiling of TRAP1-silenced breast carcinoma cells; and (ii) post-transcriptional quality control of CDK1 and MAD2, being the ubiquitination of these two proteins enhanced upon TRAP1 down-regulation. Mechanistically, TRAP1 quality control on CDK1 is crucial for its regulation of mitotic entry, since TRAP1 interacts with CDK1 and prevents CDK1 ubiquitination in cooperation with the proteasome regulatory particle TBP7, this representing the limiting factor in TRAP1 regulation of the G2-M transition. Indeed, TRAP1 silencing results in enhanced CDK1 ubiquitination, lack of nuclear translocation of CDK1/cyclin B1 complex, and increased MAD2 degradation, whereas CDK1 forced up-regulation partially rescues low cyclin B1 and MAD2 levels and G2-M transit in a TRAP1-poor background. Consistently, the CDK1 inhibitor RO-3306 is less active in a TRAP1-high background. Finally, a significant correlation was observed between TRAP1 and Ki67, CDK1 and/or MAD2 expression in breast, colorectal, and lung human tumour specimens. This study represents the first evidence that TRAP1 is relevant in the control of the complex machinery that governs cell cycle progression and mitotic entry and provides a strong rationale to regard TRAP1 as a biomarker to select tumours with deregulated cell cycle progression and thus likely poorly responsive to novel cell cycle inhibitors. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Introduction: HSP90 molecular chaperones (i.e., HSP90α, HSP90β, GRP94 and TRAP1) are potential therapeutic targets to design novel anticancer agents. However, despite numerous designed HSP90 inhibitors, most of them have failed due to unfavorable toxicity profiles and lack of specificity toward different HSP90 paralogs. Indeed, a major limitation in this field is the high structural homology between different HSP90 chaperones, which significantly limits our capacity to design paralog-specific inhibitors. Area covered: This review examines the relevance of TRAP1 in tumor development and progression, with an emphasis on its oncogenic/oncosuppressive role in specific human malignancies and its multifaceted and context-dependent functions in cancer cells. Herein, we discuss the rationale for considering TRAP1 as a potential molecular target and the strategies used to date, to achieve its compartmentalized inhibition directly in mitochondria. Expert opinion: TRAP1 targeting may represent a promising strategy for cancer therapy, based on the increasing and compelling evidence supporting TRAP1 involvement in human carcinogenesis. However, considering the complexity of TRAP1 biology, future strategies of drug discovery need to improve selectivity and specificity toward TRAP1 respect to other HSP90 paralogs. The characterization of specific human malignancies suitable for TRAP1 targeting is also mandatory.

TRAP1 is a HSP90 molecular chaperone upregulated in colorectal carcinomas and involved in control of intracellular signaling, cell cycle, apoptosis and drug resistance, stemness and bioenergetics through co-traslational regulation of a network of client proteins. Thus, the clinical significance of TRAP1 protein network was analyzed in human colorectal cancers. TRAP1 and/or its client proteins were quantified, by immunoblot analysis, in 60 surgical specimens of colorectal carcinomas at different stages and, by immunohistochemistry, in 9 colorectal adenomatous polyps, 11 in situ carcinomas and 55 metastatic colorectal tumors. TRAP1 is upregulated at the transition between low- and high-grade adenomas, in in situ carcinomas and in about 60% of human colorectal carcinomas, being downregulated only in a small cohort of tumors. The analysis of TCGA database showed that a subgroup of colorectal tumors is characterized by gain/loss of TRAP1 copy number, this correlating with its mRNA and protein expression. Interestingly, TRAP1 is co-expressed with the majority of its client proteins and hierarchical cluster analysis showed that the upregulation of TRAP1 and associated 6-protein signature (i.e., IF2α, eF1A, TBP7, MAD2, CDK1 and βCatenin) identifies a cohort of metastatic colorectal carcinomas with a significantly shorter overall survival (HR 5.4; 95% C.I. 1.1-26.6; p=0.037). Consistently, the prognostic relevance of TRAP1 was confirmed in a cohort of 55 metastatic colorectal tumors. Finally, TRAP1 positive expression and its prognostic value are more evident in left colon cancers. These data suggest that TRAP1 protein network may provide a prognostic signature in human metastatic colorectal carcinomas.

Metabolic reprogramming is one of the most frequent stress-adaptive response of cancer cells to survive environmental changes and meet increasing nutrient requirements during their growth. These modifications involve cellular bioenergetics and cross talk with surrounding microenvironment, in a dynamic network that connect different molecular processes, such as energy production, inflammatory response, and drug resistance. Even though the Warburg effect has long been considered the main metabolic feature of cancer cells, recent reports identify mitochondrial oxidative metabolism as a driving force for tumor growth in an increasing number of cellular contexts. In recent years, oxidative phosphorylation has been linked to a remodeling of inflammatory response due to autocrine or paracrine secretion of interleukines that, in turn, induces a regulation of gene expression involving, among others, molecules responsible for the onset of drug resistance. This process is especially relevant in ovarian cancer, characterized by low survival, high frequency of disease relapse and chemoresistance. Recently, the molecular chaperone TRAP1 (tumor necrosis factor-associated protein 1) has been identified as a key junction molecule in these processes in ovarian cancer: in fact, TRAP1 mediates a metabolic switch toward oxidative phosphorylation that, in turn, triggers cytokines secretion, with consequent gene expression remodeling, finally leading to cisplatin resistance and epithelial-to-mesenchymal transition in ovarian cancer models. This review summarizes how metabolism, chemoresistance, inflammation, and epithelial-to-mesenchymal transition are strictly interconnected, and how TRAP1 stays at the crossroads of these processes, thus shedding new lights on molecular networks at the basis of ovarian cancer., (© 2017 Elsevier Inc. All rights reserved.)

Ovarian cancer (OC) is the second leading cause of gynecological cancer death worldwide. Although the list of biomarkers is still growing, molecular mechanisms involved in OC development and progression remain elusive. We recently demonstrated that lower expression of the molecular chaperone TRAP1 in OC patients correlates with higher tumor grade and stage, and platinum resistance. Herein we show that TRAP1 is often deleted in high-grade serous OC patients (N=579), and that TRAP1 expression is correlated with the copy number, suggesting this could be one of the driving mechanisms for the loss of TRAP1 expression in OC. At molecular level, downregulation of TRAP1 associates with higher expression of p70S6K, a kinase frequently active in OC with emerging roles in cell migration and tumor metastasis. Indeed, TRAP1 silencing in different OC cells induces upregulation of p70S6K expression and activity, enhancement of cell motility and epithelial-mesenchymal transition (EMT). Consistently, in a large cohort of OC patients, TRAP1 expression is reduced in tumor metastases and directly correlates with the epithelial marker E-Cadherin, whereas it inversely correlates with the transcription factor Slug and the matrix metallopeptidases 2 and 9. Strikingly, pharmacological inhibition of p70S6K reverts the high motility phenotype of TRAP1 knock-down cells. However, although p70S6K inhibition or silencing reduces the expression of the transcription factors Snail and Slug, thus inducing upregulation of E-Cadherin expression, it is unable to revert EMT induced by TRAP1 silencing; furthermore, p70S6K did not show any significant correlation with EMT genes in patients, nor with overall survival or tumor stage, suggesting an independent and predominant role for TRAP1 in OC progression. Altogether, these results may provide novel approaches in OC with reduced TRAP1 expression, which could be resistant to therapeutic strategies based on the inhibition of the p70S6K pathway, with potential future intervention in OC invasion and metastasis.

Colorectal carcinoma (CRC) is a common cause of cancer-related death worldwide. Indeed, treatment failures are triggered by cancer stem cells (CSCs) that give rise to tumor repopulation upon initial remission. Thus, the role of the heat shock protein TRAP1 in stemness was investigated in CRC cell lines and human specimens, based on its involvement in colorectal carcinogenesis, through regulation of apoptosis, protein homeostasis and bioenergetics. Strikingly, co-expression between TRAP1 and stem cell markers was observed in stem cells located at the bottom of intestinal crypts and in CSCs sorted from CRC cell lines. Noteworthy, TRAP1 knockdown reduced the expression of stem cell markers and impaired colony formation, being the CSC phenotype and the anchorage-independent growth conserved in TRAP1-rich cancer cells. Consistently, the gene expression profiling of HCT116 cells showed that TRAP1 silencing results in the loss of the stem-like signature with acquisition of a more-differentiated phenotype and the downregulation of genes encoding for activating ligands and target proteins of Wnt/β-catenin pathway. Mechanistically, TRAP1 maintenance of stemness is mediated by the regulation of Wnt/β-catenin signaling, through the modulation of the expression of frizzled receptor ligands and the control of β-catenin ubiquitination/phosphorylation. Remarkably, TRAP1 is associated with higher expression of β-catenin and several Wnt/β-catenin target genes in human CRCs, thus supporting the relevance of TRAP1 regulation of β-catenin in human pathology. This study is the first demonstration that TRAP1 regulates stemness and Wnt/β-catenin pathway in CRC and provides novel landmarks in cancer biology and therapeutics.

Tumor necrosis factor receptor-associated protein 1 (TRAP1) is a heat shock protein 90 (HSP90) molecular chaperone upregulated in several human malignancies and involved in protection from apoptosis and drug resistance, cell cycle progression, cell metabolism and quality control of specific client proteins. TRAP1 role in thyroid carcinoma (TC), still unaddressed at present, was investigated by analyzing its expression in a cohort of 86 human TCs and evaluating its involvement in cancer cell survival and proliferation in vitro Indeed, TRAP1 levels progressively increased from normal peritumoral thyroid gland, to papillary TCs (PTCs), follicular variants of PTCs (FV-PTCs) and poorly differentiated TCs (PDTCs). By contrast, anaplastic thyroid tumors exhibited a dual pattern, the majority being characterized by high TRAP1 levels, while a small subgroup completely negative. Consistently with a potential involvement of TRAP1 in thyroid carcinogenesis, TRAP1 silencing resulted in increased sensitivity to paclitaxel-induced apoptosis, inhibition of cell cycle progression and attenuation of ERK signaling. Noteworthy, the inhibition of TRAP1 ATPase activity by pharmacological agents resulted in attenuation of cell proliferation, inhibition of ERK signaling and reversion of drug resistance. These data suggest that TRAP1 inhibition may be regarded as potential strategy to target specific features of human TCs, i.e., cell proliferation and resistance to apoptosis., (© 2016 Society for Endocrinology.)

Cell motility is a highly dynamic phenomenon that is essential to physiological processes such as morphogenesis, wound healing and immune response, but also involved in pathological conditions such as metastatic dissemination of cancers. The involvement of the molecular chaperone TRAP1 in the regulation of cell motility, although still controversial, has been recently investigated along with some well-characterized roles in cancer cell survival and drug resistance in several tumour types. Among different functions, TRAP1-dependent regulation of protein synthesis seems to be involved in the migratory behaviour of cancer cells and, interestingly, the expression of p70S6K, a kinase responsible for translation initiation, playing a role in cell motility, is regulated by TRAP1. In this study, we demonstrate that TRAP1 silencing enhances cell motility in vitro but compromises the ability of cells to overcome stress conditions, and that this effect is mediated by the AKT/p70S6K pathway. In fact: i) inhibition of p70S6K activity specifically reduces migration in TRAP1 knock-down cells; ii) nutrient deprivation affects p70S6K activity thereby impairing cell migration only in TRAP1-deficient cells; iii) TRAP1 regulates the expression of both AKT and p70S6K at post-transcriptional level; and iii) TRAP1 silencing modulates the expression of genes involved in cell motility and epithelial-mesenchymal transition. Notably, a correlation between TRAP1 and AKT expression is found in vivo in human colorectal tumours. These results provide new insights into TRAP1 role in the regulation of cell migration in cancer cells, tumour progression and metastatic mechanisms., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The HSP90 chaperone TRAP1 is translational regulator of BRAF synthesis/ubiquitination, since BRAF down-regulation, ERK signaling inhibition and delay of cell cycle progression occur upon TRAP1 silencing/inhibition. Since TRAP1 is upregulated in human colorectal carcinomas (CRCs) and involved in protection from apoptosis and as human BRAF-driven CRCs are poorly responsive to anticancer therapies, the relationship between TRAP1 regulation of mitochondrial apoptotic pathway and BRAF antiapoptotic signaling has been further evaluated. This study reports that BRAF cytoprotective signaling involves TRAP1-dependent inhibition of the mitochondrial apoptotic pathway. It is worth noting that BRAF and TRAP1 interact and that the activation of BRAF signaling results in enhanced TRAP1 serine-phosphorylation, a condition associated with resistance to apoptosis. Consistently, a BRAF dominant-negative mutant prevents TRAP1 serine phosphorylation and restores drug sensitivity in BRAFV600E CRC drug-resistant cells with high TRAP1 levels. In addition, TRAP1 targeting by the mitochondria-directed HSP90 chaperones inhibitor gamitrinib induces apoptosis and inhibits colony formation in BRAF-driven CRC cells. Thus, TRAP1 is a downstream effector of BRAF cytoprotective pathway in mitochondria and TRAP1 targeting may represent a novel strategy to improve the activity of proapoptotic agents in BRAF-driven CRC cells.

TNF receptor-associated protein 1 (TRAP1) is an HSP90 chaperone involved in stress protection and apoptosis in mitochondrial and extramitochondrial compartments. Remarkably, aberrant deregulation of TRAP1 function has been observed in several cancer types with potential new opportunities for therapeutic intervention in humans. Although previous studies by our group identified novel roles of TRAP1 in quality control of mitochondria-destined proteins through the attenuation of protein synthesis, molecular mechanisms are still largely unknown. To shed further light on the signaling pathways regulated by TRAP1 in the attenuation of protein synthesis, this study demonstrates that the entire pathway of cap-mediated translation is activated in cells following TRAP1 interference: consistently, expression and consequent phosphorylation of p70S6K and RSK1, two translation activating kinases, are increased upon TRAP1 silencing. Furthermore, we show that these regulatory functions affect the response to translational stress and cell migration in wound healing assays, processes involving both kinases. Notably, the regulatory mechanisms controlled by TRAP1 are conserved in colorectal cancer tissues, since an inverse correlation between TRAP1 and p70S6K expression is found in tumor tissues, thereby supporting the relevant role of TRAP1 translational regulation in vivo. Taken as a whole, these new findings candidate TRAP1 network for new anti-cancer strategies aimed at targeting the translational/quality control machinery of tumor cells., (Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Human BRAF-driven tumors are aggressive malignancies with poor clinical outcome and lack of sensitivity to therapies. TRAP1 is a HSP90 molecular chaperone deregulated in human tumors and responsible for specific features of cancer cells, i.e., protection from apoptosis, drug resistance, metabolic regulation, and protein quality control/ubiquitination. The hypothesis that TRAP1 plays a regulatory function on the BRAF pathway, arising from the observation that BRAF levels are decreased upon TRAP1 interference, was tested in human breast and colorectal carcinoma in vitro and in vivo. This study shows that TRAP1 is involved in the regulation of BRAF synthesis/ubiquitination, without affecting its stability. Indeed, BRAF synthesis is facilitated in a TRAP1-rich background, whereas increased ubiquitination occurs upon disruption of the TRAP1 network that correlates with decreased protein levels. Remarkably, BRAF downstream pathway is modulated by TRAP1 regulatory activity: indeed, TRAP1 silencing induces (i) ERK phosphorylation attenuation, (ii) cell-cycle inhibition with cell accumulation in G0-G1 and G2-M transitions, and (iii) extensive reprogramming of gene expression. Interestingly, a genome-wide profiling of TRAP1-knockdown cells identified cell growth and cell-cycle regulation as the most significant biofunctions controlled by the TRAP1 network. It is worth noting that TRAP1 regulation on BRAF is conserved in human colorectal carcinomas, with the two proteins being frequently coexpressed. Finally, the dual HSP90/TRAP1 inhibitor HSP990 showed activity against the TRAP1 network and high cytostatic potential in BRAF-mutated colorectal carcinoma cells. Therefore, this novel TRAP1 function represents an attractive therapeutic window to target dependency of BRAF-driven tumors on TRAP1 translational/quality control machinery., (©2014 American Association for Cancer Research.)

In the last decade, the identification and characterization of novel molecular mechanisms and pathways involving the heat shock protein TRAP1/HSP75 in cancers and other diseases enhanced the scientific interest. Recent reports have shown that TRAP1 stays at the crossroad of multiple crucial processes in the onset of neoplastic transformation. In fact, TRAP1: i) contributes to the tumor's switch to aerobic glycolysis through the inhibition of succinate dehydrogenase, the complex II of the mitochondrial respiratory chain; ii) is part of a pro-survival signaling pathway aimed at evading the toxic effects of oxidants and anticancer drugs and protects mitochondria against damaging stimuli via a decrease of ROS generation; iii) controls protein homeostasis through a direct involvement in the regulation of protein synthesis and protein co-translational degradation. Therefore, TRAP1 seems to be a central regulatory protein with balancing functions at the intersection of different metabolic processes during the neoplastic transformation. For this reason, it can be considered at the same time an attractive target for the development of novel anticancer strategies and a promising study model to understand the biology of tumor cells at a systemic level. This review summarizes the most recent advances in TRAP1 biology and proposes a new comprehensive view of its functions.

Adaptation to endoplasmic reticulum (ER) stress through the upregulation of the ER chaperone BiP/Grp78 favors resistance of cancer cells to anthracyclins. We recently demonstrated that the mitochondrial HSP90 chaperone TNF receptor-associated protein 1 (TRAP1) is also localized in the ER, where it is responsible for protection from ER stress and quality control on specific mitochondrial proteins contributing to its anti-apoptotic function and the regulation of the mitochondrial apoptotic pathway. Based on the evidence that Bip/Grp78 and TRAP1 are co-upregulated in about 50% of human breast carcinomas (BCs), and considering that the expression of TRAP1 is critical in favoring resistant phenotypes to different antitumor agents, we hypothesized that ER-associated TRAP1 is also favoring resistance to anthracyclins. Indeed, anthracyclins induce ER stress in BC cells and cross-resistance between ER stress agents and anthracyclins was observed in bortezomib- and anthracyclin-resistant cells. Several lines of evidence suggest a mechanistic link between the ER-stress protecting function of TRAP1 and resistance to anthracyclins: i) ER stress- and anthracyclin-resistant cell lines are characterized by the upregulation of TRAP1; ii) TRAP1 silencing in both drug-resistant cell models restored the sensitivity to bortezomib and anthracyclins; iii) the transfection of a TRAP1 deletion mutant, whose localization is restricted to the ER, in TRAP1 KD cells protected from apoptosis induced by anthracyclins; iv) the disruption of the ER-associated TRAP1/TBP7 pathway by a TBP7 dominant negative deletion mutant re-established drug sensitivity in drug-resistant cells. This process is likely mediated by the ability of TRAP1 to modulate the PERK pathway as TRAP1 KD cells failed to induce the phosphorylation of PERK in response to anthracyclins. Moreover, the downregulation of TRAP1 in combination with ER stress agents produced high cytotoxic effects in BC cells. These results suggest that ER-associated TRAP1 plays a role in protecting tumor cells against DNA damaging agents by modulating the PERK pathway.

TRAP1 is a mitochondrial antiapoptotic protein up-regulated in several human malignancies. However, recent evidences suggest that TRAP1 is also localized in the endoplasmic reticulum (ER) where it is involved in ER stress protection and protein quality control of tumor cells. Based on the mechanistic link between ER stress, protection from apoptosis and drug resistance, we questioned whether these novel roles of TRAP1 are relevant for its antiapoptotic function. Here, we show for the first time that: i) TRAP1 expression is increased in about 50% of human breast carcinomas (BC), and ii) the ER stress protecting activity of TRAP1 is conserved in human tumors since TRAP1 is co-upregulated with the ER stress marker, BiP/Grp78. Notably, ER-associated TRAP1 modulates mitochondrial apoptosis by exerting a quality control on 18 kDa Sorcin, a TRAP1 mitochondrial client protein involved in TRAP1 cytoprotective pathway. Furthermore, this TRAP1 function is relevant in favoring resistance to paclitaxel, a microtubule stabilizing/ER stress inducer agent widely used in BC therapy. Indeed, the transfection of a TRAP1 deletion mutant, whose localization is restricted to the ER, in shTRAP1 cells enhances the expression of mitochondrial Sorcin and protects from apoptosis induced by ER stress agents and paclitaxel. Furthermore, BC cells adapted to paclitaxel or ER stress inducers share common resistance mechanisms: both cell models exhibit cross-resistance to single agents and the inhibition of TRAP1 by siRNAs or gamitrinib, a mitochondria-directed HSP90 family inhibitor, in paclitaxel-resistant cells rescues the sensitivity to paclitaxel. These results support the hypothesis that ER-associated TRAP1 is responsible for an extramitochondrial control of apoptosis and, therefore, an interference of ER stress adaptation through TRAP1 inhibition outside of mitochondria may be considered a further compartment-specific molecular approach to rescue drug-resistance., (Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

We report that the mitochondrial chaperone TRAP1, which is induced in most tumor types, is required for neoplastic growth and confers transforming potential to noncancerous cells. TRAP1 binds to and inhibits succinate dehydrogenase (SDH), the complex II of the respiratory chain. The respiratory downregulation elicited by TRAP1 interaction with SDH promotes tumorigenesis by priming the succinate-dependent stabilization of the proneoplastic transcription factor HIF1α independently of hypoxic conditions. These findings provide a mechanistic clue to explain the switch to aerobic glycolysis of tumors and identify TRAP1 as a promising antineoplastic target., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Tumor necrosis factor receptor-associated protein-1 (TRAP1) is a mitochondrial (MITO) antiapoptotic heat-shock protein. The information available on the TRAP1 pathway describes just a few well-characterized functions of this protein in mitochondria. However, our group's use of mass-spectrometric analysis identified TBP7, an AAA-ATPase of the 19S proteasomal subunit, as a putative TRAP1-interacting protein. Surprisingly, TRAP1 and TBP7 colocalize in the endoplasmic reticulum (ER), as demonstrated by biochemical and confocal/electron microscopic analyses, and interact directly, as confirmed by fluorescence resonance energy transfer analysis. This is the first demonstration of TRAP1's presence in this cellular compartment. TRAP1 silencing by short-hairpin RNAs, in cells exposed to thapsigargin-induced ER stress, correlates with upregulation of BiP/Grp78, thus suggesting a role of TRAP1 in the refolding of damaged proteins and in ER stress protection. Consistently, TRAP1 and/or TBP7 interference enhanced stress-induced cell death and increased intracellular protein ubiquitination. These experiments led us to hypothesize an involvement of TRAP1 in protein quality control for mistargeted/misfolded mitochondria-destined proteins, through interaction with the regulatory proteasome protein TBP7. Remarkably, expression of specific MITO proteins decreased upon TRAP1 interference as a consequence of increased ubiquitination. The proposed TRAP1 network has an impact in vivo, as it is conserved in human colorectal cancers, is controlled by ER-localized TRAP1 interacting with TBP7 and provides a novel model of the ER-mitochondria crosstalk.

TRAP1, a mitochondrial chaperone (Hsp75) with antioxidant and antiapoptotic functions, is involved in multidrug resistance in human colorectal carcinoma cells. Through a proteomic analysis of TRAP1 coimmunoprecipitation complexes, the Ca(2+)-binding protein Sorcin was identified as a new TRAP1 interactor. This result prompted us to investigate the presence and role of Sorcin in mitochondria from human colon carcinoma cells. Using fluorescence microscopy and Western blot analysis of purified mitochondria and submitochondrial fractions, we showed the mitochondrial localization of an isoform of Sorcin with an electrophoretic motility lower than 20 kDa that specifically interacts with TRAP1. Furthermore, the effects of overexpressing or downregulating Sorcin and/or TRAP1 allowed us to demonstrate a reciprocal regulation between these two proteins and to show that their interaction is required for Sorcin mitochondrial localization and TRAP1 stability. Indeed, the depletion of TRAP1 by short hairpin RNA in colorectal carcinoma cells lowered Sorcin levels in mitochondria, whereas the depletion of Sorcin by small interfering RNA increased TRAP1 degradation. We also report several lines of evidence suggesting that intramitochondrial Sorcin plays a role in TRAP1 cytoprotection. Finally, preliminary evidence that TRAP1 and Sorcin are both implicated in multidrug resistance and are coupregulated in human colorectal carcinomas is provided. These novel findings highlight a new role for Sorcin, suggesting that some of its previously reported cytoprotective functions may be explained by involvement in mitochondrial metabolism through the TRAP1 pathway., ((c)2010 AACR.)

Background: Protein homeostasis is a highly complex network of molecular interactions governing the health and life span of the organism. Molecular chaperones, mainly heat shock proteins (HSP) and other stress-inducible proteins abundantly expressed in multiple compartments of the cell, are major modulators of protein homeostasis. TRAP1 is a mitochondrial HSP involved in protection against oxidant-induced DNA damage and apoptosis. It was recently described as a component of a mitochondrial pathway selectively up-regulated in tumor cells which antagonizes the proapoptotic activity of cyclophilin D, a mitochondrial permeability transition pore regulator, and is responsible for the maintenance of mitochondrial integrity, thus favoring cell survival. Interestingly, novel TRAP1 antagonists cause sudden collapse of mitochondrial function and selective tumor cell death, suggesting that this pathway may represent a novel molecular target to improve anticancer therapy. Preliminary data suggest that TRAP1 may be a valuable biomarker in ovarian cancers: in fact, TRAP1 levels are significantly higher in cisplatin-resistant ovarian tumors and ovarian carcinoma cell lines., Conclusions: While major advances have been made in understanding the genetics and molecular biology of cancer, given the considerable heterogeneity of ovarian cancer, the introduction of novel targeted therapies and the consequent selection of treatments based on the molecular profile of each tumor may have a major impact on the management of this malignancy and might contribute to building a new era of personalized medicine., (Copyright (c) 2009 Elsevier B.V. All rights reserved.)

TRAP1 is a component of a pro-survival mitochondrial pathway up-regulated in tumor cells. The evaluation of TRAP1 expression in 26 human colorectal carcinomas showed up-regulation in 17/26 tumors. Accordingly, TRAP1 levels were increased in HT-29 colorectal carcinoma cells resistant to 5-fluorouracil, oxaliplatin and irinotecan. Thus, we investigated the role of TRAP1 in multi-drug resistance in human colorectal cancer. Interestingly, TRAP1 overexpression leads to 5-fluorouracil-, oxaliplatin- and irinotecan-resistant phenotypes in different neoplastic cells. Conversely, the inhibition of TRAP1 activity by TRAP1 ATPase antagonist, shepherdin, increased the sensitivity to oxaliplatin and irinotecan in colorectal carcinoma cells resistant to the single agents. These results suggest that the increased expression of TRAP1 could be part of a pro-survival pathway responsible for multi-drug resistance.

TRAP-1 is a mitochondrial heat shock protein (HSP), recently identified in Saos-2 osteosarcoma cells adapted to mild oxidative stress induced by diethylmaleate (DEM). TRAP-1 mRNA expression is increased in DEM-adapted cells as well as in tumor cells resistant to 5-fluorouracil and to platin derivatives. Since a strong decrease of TRAP-1 protein levels, upon cisplatin treatment, is observed only in controls but not in the DEM-adapted counterpart, a possible role for this protein in the development of resistant phenotypes could be hypothesized. To characterize the protective role of TRAP-1 against oxidative stress and apoptosis, stable transfectants were generated and characterized for their response to different stress types. These stable clones expressing constitutively high TRAP-1 levels: (i) are more resistant to H2O2-induced DNA damage and to apoptosis by cisplatin; (ii) contain higher reduced glutathione (GSH) levels than control cells; and (iii) do not release the apoptosis-inducing factor into the nucleus upon cisplatin treatment. Furthermore, high TRAP-1 levels interfere with caspase 3 activation. These results confirm the anti-apoptotic role of TRAP-1, and suggest that increased expression of this mitochondrial HSP in DEM-adapted and chemoresistant cells could be part of a pro-survival signaling pathway aimed to evade toxic effects of oxidants and anticancer drugs.