Implications of endoplasmic reticulum stress, the ...

Review

1.

Introduction

2.

Targeting the ERS, the UPR and apoptosis for cancer therapy

3.

Conclusion

4.

Expert opinion

Implications of endoplasmic reticulum stress, the unfolded protein response and apoptosis for molecular cancer therapy. Part II: targeting cell cycle events, caspases, NF-kB and the proteasome Donavon C Hiss† & Gary A Gabriels¶ †University

of the Western Cape, Department of Medical BioSciences, Molecular Oncology Research Programme, Bellville, 7535, South Africa; ¶University of Cape Town, Observatory, Division of Clinical Pharmacology, Department of Medicine, 7925, South Africa

d Lt ion K ut a U trib rm is o f D In i al 9 c 0 er 20 m © m t o h ig or C r py o ale C S or f t No

Background: Endoplasmic reticulum stress (ERS), the unfolded protein response (UPR) and apoptosis signal transduction pathways are fundamental to normal cellular homeostasis and survival, but are exploited by cancer , ad activation of cells to promote the cancer phenotype. Objective: Collateral o l wn e. or apoptosis, ERS and UPR role players impact on cell growth, cell cycle do arrest s n a al u aggressiveness genomic stability, tumour initiation and progression, n s c stumour o r e er and drug resistance. An understanding of these affords promising us processes d rp o e f prospects for specific cancer drug targeting of the ERS, UPR and apoptotic s ri py thoII)cobrings pathways. Method: This review (Part IIuof forward the latest devel. A ingle d opments relevant to the molecular connections among cell cycle regulators, e bit t a s i h caspases, NF-κB, and the proteasome o in with ERS and UPR signalling cascades, pr pr einduction, their functions in apoptosis apoptosis resistance and oncogenesis, s nd d u ew acan be exploited for targeted cancer therapy. and how these relationships e ris , vi Conclusion: Overall, tho ERS, ay the UPR and apoptosis signalling cascades (the u a ispl n molecular therapeutic targets) and the development of drugs that attack d U these targets signify a success story in cancer drug discovery, but a more reductionist approach is necessary to determine the precise molecular switches that turn on antiapoptotic and pro-apoptotic programmes. Keywords: anticancer drugs, apoptosis, cancer, cancer drug discovery, caspases, cyclin-dependent kinase inhibitors, cyclin-dependent kinases, cyclins, ERS, inhibitors-of-apoptosis proteins, NF-κB, oncogenes, proteasome inhibitors, signal transduction, tumour suppressor genes, UPR, UPS Expert Opin. Drug Discov. (2009) 4(9):907-921

1.

Introduction

The physiological and pathological significance of the endoplasmic reticulum stress response (ERS) and the unfolded protein response (UPR) signal transduction pathways are well documented (see Review Part I of II for more details). ERS and UPR pathways have long been recognised as molecular targets in the search for specific therapeutic rationales that can be applied effectively in various malignancies [1-20]. The UPR and the endoplasmic reticulum-associated degradation pathway constitute a multiplex signalling system in which several collateral mechanisms purge the cell of toxic unfolded protein aggregates. Thus, inhibition 10.1517/17460440903055032 © 2009 Informa UK Ltd ISSN 1746-0441 All rights reserved: reproduction in whole or in part not permitted

907

Implications of endoplasmic reticulum stress, the unfolded protein response and apoptosis for molecular cancer therapy

of protein synthesis or alteration of the degradation and folding capacity of the endoplasmic reticulum (ER) affords both a powerful and plausible incentive to study the molecular players that dictate the efficiency of the UPR to relieve ER stress and to ensure cell survival. The potential application of observations that cell cycle arrest, growth inhibition as well as induction of apoptosis in various tumour cell types can be initiated by perturbation of the UPR is emerging for the development of novel proteasome inhibitors as anticancer drugs [19-24]. The role of the UPR in development, differentiation and the cellular pathways leading to cell survival (cell proliferation and self-renewal to maintain normal cell numbers), apoptosis (cessation of cell proliferation and initiation of cell death) or oncogenesis (uncontrolled cell proliferation and tumour formation) is perhaps best illustrated by current evidence implicating ubiquitin-mediated proteolysis as the chief modulator of haematopoietic stem-cell regulators that control the fate of myeloid and erythroid cells [25,26]. More specifically, culin 4A has been identified as the ubiquitin ligase that controls the proliferation, differentiation, maturation and destiny of haematopoietic progenitor cells. Lack of culin 4A causes DNA damage and apoptosis of these rapidly dividing cells, leading to bone marrow failure and insufficient numbers of mature blood cells, thus, culminating in leukaemia, anaemia and other generalised sequelae of myelosuppression [26]. Clearly, delineation of the molecular mechanisms that drive normal haematopoietic stem-cell differentiation and functioning of the UPR in normal and malignant haematopoiesis may prove significant in the development of new treatments for haematological malignancies and other cancers [27]. This review (Part II of II) provides new insights into the molecular connections of cell cycle regulators, caspases, NF-κB, and the proteasome with ERS and UPR responses, their functions in apoptosis induction, apoptosis resistance and oncogenesis, and how these relationships can be exploited for targeted cancer therapy. 2. Targeting the ERS, the UPR and apoptosis for cancer therapy 2.1

Cell cycle events

Induction of ER stress and the UPR is generally associated with diminished cell proliferation due to G1 arrest through loss of cyclins D1 and G1 [28-31]. ER stress, the UPR, apoptosis and cell cycle control signal transduction pathways are fundamental to cell kinetics and tissue homeostasis [32], but the precision of these pathways and their integration are compromised during sustained exposure to cytotoxic agents, and further deregulated in cancer cells. Cell-cycle regulatory serine/threonine kinases (cyclin-dependent kinases; CDKs), CDK inhibitors (e.g., p21 and p27) and NF-κB are all subject to the UPR proteolytic machinery that impacts on cell growth, cell cycle arrest or apoptosis, genomic stability, tumour initiation and progression, tumour aggressiveness and drug resistance [2,14,33-47]. Several downstream genes of p53, such as p21, cyclin G and GADD153/CHOP (see Review Part I 908

of II for more details) regulate various aspects of cell growth cascades and are rapidly upregulated after chemo- and radiation-induced DNA damage. The induction of p21 mediates p53-dependent G1 arrest through its inhibitory effects on CDKs required for S-phase entry [30,48]. Degradation of cyclins by the ubiquitin-proteasome system (UPS) is vital for the control of cell cycle progression and deregulation of the UPR might result in uncontrolled cell proliferation, genomic instability and cancer [49]. CDK inhibitors regulate the assembly and activity of the cyclin-CDK kinase complexes and are broadly classified into two families: INK4 and Cip/Kip (CDK inhibitory protein/kinase inhibitory protein). The INK4 family (p16INK4A, p15INK4B, p18INK4C and p19 INK4D) preferentially bind and block CDK4/6, whereas the Cip/Kip family (p21cip, p27kip1 and p57kip2) have a wide-ranging specificity in inhibiting CDK activity [46,50]. Cell cycle transition from the G1 to the S phase epitomises the crossroad or decision point (restriction point) where cells either continue to divide and complete the cell cycle or withdraw from the cycle. Mutations that lead to gain-of-function (oncogenes) or loss-of-function (tumour suppressor genes) exert their effects at this critical juncture. Control of the G1/S restriction point is mediated by retinoblastoma tumour suppressor protein (Rb or pRb) through complex interactions with cyclins and CDKs. Active nonphosphorylated Rb protein impedes transit through the restriction point, whereas phosphorylation inactivates Rb, opposing its inhibitory function and facilitating cell cycle traverse [51]. Recently, a systems biology dynamical model of mammalian G1 cell cycle progression has been conceptualised for the development of therapeutics that target cell-cycle events. This mathematical model incorporates essential tenets of G1 cell-cycle progression and relies profoundly on growth factor-induced upregulation of cyclin D:Cdk4/6 complex activity to partially inactivate pRb by phosphorylation and to sequester p27Kip1, and to activate cyclin E:Cdk2 complexes that in turn inactivate pRb, which oscillates between an active hypophosphorylated conformation associated with E2F transcription factors in early G1 phase and an inactive hyperphosphorylated conformation in late G1, S and G2/M phases [52]. This model illustrates significant evolution towards the identification and application of radically adjusted methodologies to profile and customise cell-cycle targeted chemotherapy. Moreover, the potential use of antibodies to downregulate cyclin E overexpression in cancers has proved progressively more feasible in the face of the spiralling demand to devise new generation antibody-dependent tumour cytotoxicity enhancement [53]. Another move forward to perk up tumour selectivity is the development of subsequent clinical trials with the multi-target tumour growth inhibitor™ (MTGI™, ZK 304709), specific for CDK-1 and CDK-2 as well as VEGFRs 1, 2, 3 and platelet-derived growth factor receptor. The efficacy of ZK 304709 is based on the maxim that simultaneous inhibition of cell-cycle progression and neoangiogenesis might produce superior cancer eradication through its ability to inhibit vascular permeability, cell cycle

Expert Opin. Drug Discov. (2009) 4(9)

Hiss & Gabriels

molecular targets, as well as tumour growth in human xenografts [54,55]. Multi-target anticancer agents such as ZK 304709 portray an encouraging new therapeutic approach for targeting the tumour microenvironment, the site where many events and interactions such as adhesion, survival, proteolysis, migration, immune escape mechanisms, lymph-/angiogenesis, and metastasis or homing on target organs occur [56]. Deregulated expression of cyclin D1 and cyclin E as well as CDK4 and CDK2 has been identified in human oesophageal squamous cell carcinoma, and shows a logical relationship to tumour staging, metastases, and poor response to chemotherapy, and decreased overall survival [57,58]. Likewise, the expression of cyclin D1 correlates with growth phenotypes of MCF7erbB2 drug- sensitive and -resistant breast cancer cells treated with 2′-deoxy-5-fluorodeoxyuridine or 1-β-D-arabinofuranosylcytosine, and may serve as a marker to evaluate chemosensitivity during cell cycle inhibition [59]. CDK inhibitors have been evaluated for their efficacy alone or in combination with standard anticancer drugs [60]. Flavopiridol, a synthetic flavone and an inhibitor of several CDKs with antiproliferative and pro-apoptotic properties, has been shown in clinical trials to cause tumour regression [61], and exhibits sequence-dependent cytotoxic synergy with docetaxel in preclinical gastric cancer models [62]. Differential expression of the CDK inhibitor p27kip1 and the SCF E3-ubiquitin ligase that controls its stability (Skp2) in tumours, including prostate cancer, colon carcinoma, and small cell lung cancer, have been shown to correlate with disease prognosis [63-65]. Because p27kip1 is a tumour suppressor protein, its expression is downregulated and degradation upregulated in most human cancers. Application of this knowledge has led to the identification of yet another proteasome inhibitor, argyrin A, which exerts its anticancer activity by blocking proteasomal degradation of p27kip1, thus stabilising p27kip1 levels and concomitantly inhibiting the acquired resistance to argyrin A attributed to loss of p27kip1 expression. The clinical development and use of argyrin A may provide a targeted approach to the treatment of many cancers because of its specificity for p27kip1, a property not shared by the widely acclaimed proteasome inhibitor, bortezomib. Notwithstanding, the combination of argyrin A and bortezomib still needs to be explored for synergistic efficacy with a view to establishing multipronged and enhanced proteasome inhibition in accord with the ethic of tolerable dose levels [65-67]. Because p21 and p27 are downstream effectors of the EGFR cascade (a tyrosine kinase pathway deregulated in many cancers, especially breast carcinoma) that communicate proliferative signals, cross-talk between the proteasome and the EGFR pathway is also extensively being investigated, both in terms of basic preclinical and clinical studies [2,14]. Clearly, regulation by the UPS (through post-translational ubiquitylation and degradation by the 26S proteasome) of CDKs, kinase inhibitors and other protein kinases involved in growth-factor-mediated signal transduction pathways and critical cell cycle points is

central to probing cell transformation and cancer progression, and the specific design of agents that target these molecular processes [68,69]. 2.2

Caspases

The caspases represent a family of cysteine proteases that are important molecular players in apoptosis signalling [70-73]. Caspases have several roles in cell death mechanisms, including initiator, activator and effector functions [74], as well as caspasedependent alterations of the UPR [75]. Apoptosis is regulated by the intrinsic and extrinsic pathways that connect the three proximal arms of the UPR (activating transcription factor-6 [ATF6], protein kinase RNA-like ER kinase [PERK], inositol-requiring endoribonuclease protein-1 [IRE1]). The intrinsic pathway responds to ERS and DNA damage, and must maintain the balance between pro-apoptotic BH3-only proteins (e.g., Bad, Bak and Bax) and antiapoptopic proteins (e.g., Bcl-2 proteins). ERS triggers conformational changes such as oligomerisation of Bak and Bax localised to the ER membrane, thus, causing Ca2+ efflux from the ER lumen and activation of cytosolic calpain. Calpain catalyses the activation of ER-resident procaspase-12 to caspase-12, which, in turn, sets off the caspase cascade by cleavage and activation of procaspase-9 and procaspase-3. The extrinsic pathway is activated by ERS and is characterised by the formation of a heterotrimeric complex among IRE1, TNF receptor-associated factor 2 (TRAF2) and apoptosis signal-regulating kinase 1 (ASK1), and activation of JNK and cell death. Moreover, ATF6 and PERK signalling cascades converge on the downstream effector growth arrest and DNA damage-inducible protein 153 (GADD153)/CHOP that blocks antiapoptotic Bcl-2, and, therefore, results in growth arrest and cell death [76]. Mitochondrial and ERS-induced apoptosis pathways display parallel activation of caspase-12 and caspase-9 cascades [77,78]. Bromovulone III (a marine prostanoid extract), for example, showed anticancer efficacy against human hepatocellular carcinoma cells by inducing ERS (as evidenced by activation of calpain, caspase-12 and GADD153/CHOP) and ER-mitochondrial cross-talk [78,79]. In a number of tumours, β-lapachone (an o-naphthoquinone) has been found to induce apoptosis, but in human prostate carcinoma, the pro-apoptotic effects of this compound (procaspase-12 activation, phosphorylation of p38, ERK, JNK, and activation of executioner caspases, caspase-7 and calpain) have been expressly linked with ERS indicators, such as raised cytosolic Ca2+ levels, upregulation of GRP78/BiP and GADD153/CHOP [80]. Because ER stress can induce caspase expression [81-84], caspases are indicated in apoptotic intervention and considered to be important anticancer targets for drug development [85-94]. In human cancers, downregulation of caspases suppresses apoptosis and promotes other attendant components of neoplasia, such as assertive clonal expansion, escape from immune surveillance and attack, metastasis, and resistance to chemo- and radiation therapy. Mammalian endogenous inhibitor-of-apoptosis proteins (IAPs), such as XIAP, cIAP-1, cIAP-2, NAIP, ML-IAP, ILP-2 and survivin,


909


block caspases by binding to them and tagging them (by ubiquitylation) for UPS-mediated degradation [73,95]. IAPs function as decoys and compete with specific caspase targets to stall downstream events that are indispensable for the execution of cell death. Thus, IAPs overexpressed in cancers short-circuit caspase-dependent signalling pathways that converge, through the UPR proximal arms, on apoptosis [96]. The antiapoptotic ability of IAPs is exemplified by the oncogene, cIAP-1, a member of the IAP protein family and a RING finger ubiquitin E3-ligase, which is upregulated in many cancers. cIAP-1 catalyses ubiquitylation of Max-dimerisation protein-1 (Mad1), a cellular antagonist of c-Myc, resulting in its rapid degradation by the 26S proteasome. Thus, cIAP-1-induced reduction of Max-dimerisation protein-1 levels drives c-Mycassociated amplification of tumour cell proliferation [97]. The 26S proteasome, a multi-catalytic threonine protease complex responsible for intracellular protein turnover in eukaryotic cells is a major chemotherapeutic target [68,98,99]. Clinical prognosis in patients with nodal diffuse large B-cell lymphomas (DLBCLs), for instance, has been correlated with IAPs, such as X-linked inhibitor of apoptosis protein (XIAP) that arrests apoptosis by inhibiting active caspase-3, caspase-7 and caspase-9. The utilisation of a small molecule antagonist of XIAP (1396-12) against lymphoma cells derived from patients with DLBCL attenuated caspase-3 inhibition and induced apoptosis in 16 of the 20 DLBCL samples analysed. XIAP antagonism was effective in both chemotherapy-refractory and -responsive DLBCL cells, the latter showing raised expression of XIAP, decreased antiapoptotic Bcl-2 levels and constitutive caspase-9 activation [100]. In the broader context, the studies described above exemplify the use of XIAP antagonists as biological markers to predict therapeutic outcome in cancer patients with high expression levels of IAPs. In another advance on DLBCL cells, it was demonstrated that small molecule Bcl-2 antagonists potentiated the tumouricidal effects of bortezomib through mitochondrial mechanisms, enhanced JNK activation and ER stress induction (e.g., eIF2α phosphorylation, activation of caspases-2 and -4, and GRP78/BiP upregulation) [101]. The GOLF regimen (gemcitabine, oxaliplatin, leucovorin and 5-fluorouracil), highly rated for its efficacy and safety profile in metastatic colorectal carcinoma, has been shown to induce apoptosis in human colon cancer cells by poly (ADP-ribose) polymerase cleavage, caspase-9 and caspase-3 activation, UPR-regulated reduction in the expression of the upstream activators Raf-1 and Akt, and inactivation of the multi-chaperone complex [102]. It is widely accepted that apoptosis resistance is conducive to oncogenesis and the establishment of a resistant cancer phenotype that leads to failure of cancer chemotherapy. A recent study thoroughly merited this view in that the cytotoxic drugs methotrexate and 5-fluorouracil sensitised a broad panel of resistant tumour cell lines and TNF-related apoptosis-inducing ligand (TRAIL)-resistant primary acute leukaemia cells for induction of apoptosis by p53-mediated upregulation of caspase-8. Upregulation of caspase-8 abolished 910

TRAIL-induced cellular proliferation and shifted sensitivity toward TRAIL-induced apoptosis, as confirmed by short hairpin RNA (shRNA)-knockdown or silencing of both p53 and caspase-8 [84]. Similar results have been published for the activation of caspase-9 and caspase-3 in drug resistant human NSCLC cells, strongly suggesting that delineation of apoptosis defects may be key to screening for prognostic markers and to designing novel apoptogenic (aptoptosis-inducing) or caspase-activation therapeutic strategies [103]. A recent report specifically focused on this proof-of-principle in a clinical trial in paediatric acute myeloid leukaemia patients and, as in the case with childhood acute lymphoblastic leukaemia, asserted that intact apoptosis signalling is indicative of favourable treatment outcome, as validated by the measurement of two apoptogenic events, cytochrome c release and caspase-3 activation [104]. This study, therefore, provided convincing evidence that in patients with good prognosis, cytochrome c release was caspase-dependent and correlated with activated caspase-3 (caspase-dependent cytochrome c-related activation of caspase-3 being the purported parameter). Interestingly also, proteasome inhibition by bortezomib significantly sensitised prostate cancer cells to apoptosis initiated by Fas ligand or TRAIL and upregulated caspase-8 potency [12]. These findings indicate the spurt of research efforts that target dysregulated apoptosis as a model for efficacy-based cancer therapies [71,105,106]. 2.3

NF-κB

The UPR is an integrated genomic response to ER stress and critical for the degradation of proteins involved in cell cycle regulation, apoptosis and angiogenesis. Tumour cells are generally resistant to apoptosis and are able to confound UPR-mediated regulation of steady-state protein levels, thus, resulting in gain-of-function (overexpression of proto-oncogenes and their mutant overactive forms, oncogenes) and loss-of-function (decreased expression of tumour suppressor genes) that promote tumourigenesis and manifestation of the dominant cancer phenotype, which confer a hyperproliferative signature, enhanced drug resistance and metastatic potential [17,71,107-111]. The acquisition of resistance to apoptosis can occur by up- or downregulation, through the UPR, of many of the apoptotic role players. Besides GADD153/CHOP and p53 (see Review Part I of II for more details), the activity of another transcription factor, namely, NF-κB, which is constitutively expressed in many cancer cells, is also modulated by the ERS and the UPR [17,19,112-115]. NF-κB activation is mediated by ER stress through the IRE1-TNF receptor-associated factor 2 pathway and JNK activation [116]. NF-κB is of special interest in tumour immunology because its activation along with that of other transcription factors such as signal transducer and activator of transcription (STAT1 and STAT3) is generally associated with signalling pathways of pro-inflammatory cytokines such as TNF-α, IFN-γ, IL-1, IL-4, IL-6, and GM-CSF [117]. In tumour cells, NF- κ B and STAT3


Hiss & Gabriels

promote cell growth and survival by inducing pro-inflammatory target genes that stall apoptosis and antiapoptotic proteins (Bcl-2, Bcl-XL, cIAP, c-Flip), whereas STAT1 functions as an antioncogene by mediating apoptosis through p53 [117]. The activation of NF-κB is dependent on the 26S proteasome [118] and involves several proteins, including the zinc finger protein A20 known to exert dual, but opposing UPS effects – successive de-ubiquitylation and ubiquitylation of the TNF receptor-interacting protein (RIP) – thereby targeting RIP to proteasomal degradation [119,120]. High expression levels of NF-κB in human cancers, including multiple myeloma, breast cancer and squamous cell carcinoma, may occur as a result of TNF-α- or anticancer drug-potentiated phosphorylation and ubiquitylation, and subsequent degradation of I-κB (an endogenous inhibitor of NF-κB) that propels nuclear translocation of NF-κB and NF-κB-induced gene transcription and inhibition of apoptosis [118,121-125]. NF-κB induces the transactivation of numerous target genes that inhibit apoptosis and promote cell division [117]. Thus, overexpression of NF-κB in many cancers correlates with the induction of antiapoptotic genes such as c-Flip, cIAP-1, cIAP-2 and XIAP that block the caspase cascade or antiapoptotic members of the Bcl-2 family that prevent mitochondrial cytochrome c release, a marker of apoptosis [113,118,126]. The activation of NF-κB in response to chemo- or radiation therapy often results in acquired apoptosis resistance that presents a major obstacle to successful treatment outcome. The pleiotropic effects of NF-κB on cell proliferation is perplexing because it stimulates the expression of both inducers and repressors of cell cycle progression, for example, NF-κB upregulates cyclin D or E20 as well as p21Cip1 expression [113,118]. In many tumours, transcriptionally-induced overexpression of Bcl-2 is linked to chemo- and radiation resistance. In a number of cancer cell lines, the 26S proteasome inhibitor, bortezomib, has been shown to inhibit NF-κB acivity and to concomitantly decrease antiapoptotic Bcl-2 levels [127]. The anilinopyrimidine derivative, AS602868, a pharmaceutical inhibitor of I-κB kinase 1 (IKK1), has been evaluated for its anticancer effects, either as a single agent or in combination with suboptimal doses of melphalan or bortezomib, in a human multiple myeloma cell line and patient-derived primary myeloma cells. AS602868 synergistically inhibited cell proliferation in the multiple myeloma cell line and induced apoptosis of primary myeloma cells without affecting the survival of bone marrow mononuclear cells (CD138-) coincubated with primary multiple myeloma (CD138+) cells. These findings reveal a significant role for NF-κB in the pathogenesis and treatment of multiple myeloma [128]. In a similar study, the combination of bortezomib with the histone deacetylase inhibitor, suberoylanilide hydroxamic acid, diminished NF-κB signalling in mantle cell lymphoma [129]. Because mantle cell lymphoma is a highly incurable B cell lymphoma, these observations may have applications in cancer therapy regimens that combine proteasome inhibitors with

agents that promote histone acetylation. In addition, the use of a novel adenoviral expression system to deliver an RNA aptamer that selectively targets and inhibits NF-κB activation in a human NSCLC cell line and a lung tumour xenograft model, blocked doxorubicin resistance and angiogenesis induced by NF-κB through the hypoxia-inducible factor1α/VEGF pathway [130]. Moreover, inactivation of the NF-κB pathway can be induced by caspase-dependent cleavage of IKK1 and NEMO (NF-κB essential modulator), which are needed to transduce NF-κB activation signals. NEMO, but not IKK1, is thought to be essential for the antiapoptotic (pro-survival) effects of NF-κB because inactivation of NEMO caused marked downregulation of antiapoptotic NF-κB target genes encoding cIAP-1 and cIAP-2 (caspase inhibitors) or adaptors of TNF-α [131]. It is obvious that NF-κB is a prime accomplice in oncogenesis and the protection of cancer cells from apoptosis [132], and, therefore, has become a major target for anti-inflammatory and anticancer drug development [133-135]. Besides this, NF-κB is also known for its antagonistic effects on p53 [136], an interaction that warrants further investigation. 2.4

The proteasome

Compounds that interfere with proteasome function exert their activities through a variety of mechanisms, including inhibition of cellular proliferation, induction of apoptosis, synergism with chemo-, radiation, and immune therapy, and reversal of antitumour drug resistance [13,18,20]. In recent years, the proteasome has become a recognised site in the quest for identifying and probing potential avenues of new anticancer drug targets in both solid and haematologic tumours [2,4-7,9,14,20,45,137-139]. Epoxomicin, one of the earliest documented proteasome inhibitors that also exhibited evidence of in vivo anti-inflammatory activity [20,140], has recently been shown to activate the transcription of both PUMA (p53 upregulated modulator of apoptosis) and Bim in human colon cancer cells [137]. Because upregulation of PUMA and Bim are also typical features of ER stress-induced apoptosis, such cellular responses underscore the weight that should be accorded to the p53/PUMA pathway in determining tumour cell sensitivity to proteasome inhibitors and the design of pro-apoptotic anticancer agents [19,141]. However, one of the most remarkable advances in proteasome inhibition has been the approval of bortezomib (Velcade®, Millennium Pharmaceuticals, Cambridge, MA, USA) for the treatment of multiple myeloma patients [22,142-146]. Bortezomib is a prototype proteasome inhibitor with anticancer activity either as a single drug or in combination with other antitumour agents [1,2,45,129,143,147-150]. Bortezomib’s antineoplastic, proapoptotic and antiangiogenic activities are closely linked to apoptosis through its ability to inhibit the NF-κB pathway, its pleiotropic effects on p53, p21waf1/cip1 and p27kip1, apoptotic proteins Bid, Bim, Bax, Bak, Bik, NOXA, caveolin-1 and I-κB (which blocks NF-κB-mediated induction of pro-survival signal transduction pathways), and activation of the caspase


911


cascade [45,117,148,151-155]. NPI-0052, another novel proteasome inhibitor that differs from bortezomib in chemical structure and mechanism of action is undergoing validation for preclinical anticancer efficacy. In multiple myeloma cells, NPI-0052 has well-defined synergistic pro-apoptotic effects in combination with bortezomib, including, activation of caspase-3, caspase-8, caspase-9 and poly (ADP-ribose) polymerase, stimulation of ERS and JNK, inhibition of cell migration and angiogenesis, and impairment of NF-κB signalling [156,157]. In multiple myeloma, substantial amounts of antibodies are produced causing considerable ERS which, in turn, calls on the UPR to clear the large protein burden [158]. Combination of bortezomib and the death receptor ligand, TRAIL, provoked a synergistic apoptotic response in prostate and colon cancer cell lines. In a number of cell lines, bortezomib seems to increase Bik and Bim, but not PUMA, Bid, Bax, Bak, Bcl-2 and Bcl-xL. The enhancement of TRAILinduced apoptotis by bortezomib correlates with inhibition of proteasome-dependent cleavage of Bik and Bim, because silencing of their genes in mouse embryo fibroblasts renders the host cells resistant to bortezomib cytotoxicity. This effect was further corroborated by abrogation of synergy between bortezomib and TRAIL in human prostate cancer cells when Bik and Bim were downregulated by RNA interference and a deficiency of APAF-1, which functions downstream of Bcl-2 [151]. Bortezomib also induces apoptosis in primary Waldenström’s macroglobulinaemia (WM) cells and WM cell lines [159]. Furthermore, oblimersen sodium (G3139), an antisense oligonucleotide designed to specifically target the first 6 codons of human Bcl-2 mRNA, demonstrated proof-ofprinciple of an antisense effect in human tumours and an apoptosis-modulating strategy by downregulating Bcl-2 protein, synergising with many cytotoxic and immunotherapeutic agents against a variety of cancers, including WM, acute myeloid leukaemia, chronic lymphocytic leukaemia, multiple myeloma, malignant melanoma, NSCLC, non-Hodgkin’s lymphoma, gastric, colon, bladder and Merkel cell cancers, as well as hormone-refractory prostate cancer [160,161]. Because increased ER stress leads to a pro-apoptotic and terminal UPR in WM cells, these molecular studies, along with those using the antioxidant resveratrol (3,4′,5-tri-hydroxy-trans-stilbene) and inhibitors of ubiquitin E3-ligase that selectively modulate the expression of receptors, growth factors and transcription factors essential to the growth, survival and spread of tumours, present a useful context for rational design of the next generation of combination apoptosis-induction and proteasome-inhibition therapies for WM and other malignancies [162-167]. Synergistic interaction of bortezomib with TNF-α in an experimental mouse colon carcinoma model illustrates the advantage of collateral sensitivity of tumour cells to combination chemotherapeutic regimens over the limited efficacy of single-agent exposure [1]. In tumour-bearing mice, the mechanistic relation of the synergism between bortezomib and TNF-α has been confirmed to include caspase-3 and caspase-12 cleavage, p53 accumulation, increased SAPK/JNK 912

phosphorylation, upregulation of GRP78/BiP, PDI and calnexin, all chronic ERS and pro-apoptotic processes that substantially suppressed tumour growth and increased animal survival rate. By contrast, TNF-α counteracted bortezomib-induced upregulation of Hsp27 [1]. Such bifurcate observations may provide a better insight into synergistic or antagonistic proteasome-anticancer drug discovery rationales. The constitutive expression of Hsp27 in many tumours together with its antiapoptotic and pro-survival roles that protect cancer cells against ERS and anticancer drug-induced genotoxic stress must, as logical determinants, be incorporated into future experimental or clinical blueprints for proteasomebased cancer chemotherapies [168-171]. Another important consideration is the development of clinical resistance to bortezomib, which may present a significant impediment to its efficacy as an anticancer drug [138]. However, optimism abounds that studies on bortezomib’s interaction with the multi-drug transporter (P-glycoprotein) and the multi-drug resistance protein 1 may allay our lack of insight into the molecular events involved in the expression of resistance and cross-resistance to bortezomib in tumours with persistent high incidence and mortality rates [138]. Likewise, the use of biological response modifiers or resistance sensitisers in combination with bortezomib to modulate proteasome function and enhance degradation of misfolded P-glycoprotein may be invaluable. Noteworthy accounts of experimental mani pulations to reverse P-glycoprotein-mediated drug resistance have appeared steadily and may prove advantageous in this regard [138,172-174]. Clinical experience with bortezomib alone and in combination with other oncolytic drugs in haematologic malignancies has been amply documented [22,175]. In a Phase II clinical trial with relapsed multiple myeloma patients, the combination of bortezomib (Velcade®) with intermediate-dose dexamethasone and continuous low-dose oral cyclophosphamide yielded a 90% efficacy rate (median event-free survival of 12 months and median overall survival of 22 months), although adverse events such as leukopaenia, infection, herpes zoster, thrombocytopaenia, neuropathy and fatigue were observed in at least 10% of patients. Compared to bortezomib monotherapy, the bortezomib/dexamethasone/cyclophosphamide combination regimen with obligatory antiviral prophylaxis improved the response rate and event-free survival rate in relapsed multiple myeloma patients [176]. Bortezomib displayed synergy with anticancer drugs such as 5-fluorouracil, paclitaxel and doxorubicin in human gastric cancer cell lines [147]. Similarly, the combination of doxorubicin and melphalan triggered cell death in refractory multiple myeloma cells, independent of the antiapoptotic protein myeloid cell leukaemia-1 (Mcl-1) whose overexpression in these tumour cells protects them from chemotherapy-induced apoptosis [177]. Because bortezomib elicited both downregulation of Mcl-1 and its caspase-dependent degradation, the combination of bortezomib with doxorubicin and melphalan suggests an alternative course to overcome relapse in multiple myeloma


Hiss & Gabriels

patients [177]. Interestingly, a recent study purported that cytoreductive chemotherapy using bortezomib in tandem with the BRC regimen (cyclophosphamide and rituximab) may offer improved efficacy in aggressive B-cell lymphoma with poor clinical outcome [178]. Regardless of the advances in proteasome cancer chemotherapeutic modalities, the predictable apprehension about acquired resistance following unabated exposure to bortezomib calls for prudence [10,179], especially from the perspective of amplified de novo biogenesis of 19S-20S-19S proteasomes boosted by increased expression of the proteasome maturation protein (POMP) and antiapoptotic Hsp27 protein, as well as impaired pro-apoptotic p53 protein upregulation and stabilisation in human Burkitt lymphoma cells [180]. Thus, the aforementioned events connote an adaptive proteasome response to continuous bortezomib treatment to confer a survival advantage or ability to assert a hyper proliferative and apoptosis-resistant phenotype in cells confronted with severe proteasome stress [180]. A recent study showed that combination of bortezomib with recombinant adeno-associated virus type 2- mediated p53 gene transfer or docetaxel and pemetrexed produced significant synergistic inhibition of cellular proliferation in p53-positive NSCLC cells, an effect that will have to be viewed as a contingency in clinical trials using bortezomib, docetaxel and pemetrexed [3]. Frontline targeted oncologic therapies are expanding with the development of novel proteasome inhibitors such as NPI-0052 and carfilzomib, which may engender elevated remission rates in multiple myeloma [5]. 3.

Conclusion

Cell cycle-dependent kinases (CDKs) and CDK inhibitors such as p21 and p27 regulate various aspects of cell growth cascades. Because levels of CDKs and CDK inhibitors are regulated by the UPR, they are of particular interest in targeting the apoptotic (cell death) and cell survival pathways. Moreover, alterations in levels and function of these cell cycle regulators may provide important cues with regard to cancer pathogenesis, clinical diagnosis and prognosis [46]. The G1 to the S phase is the checkpoint where cells are either restricted or permitted to continue their journey through the cell cycle. Many oncogenes and tumour suppressor genes exert their effects at this critical juncture. The tumour suppressor protein, Rb, mediates growth control of the G1/S decision point by complex interactions with cyclins and CDKs. Active nonphosphorylated Rb blocks transit of cells through the restriction point, whereas inactive phosphorylated Rb facilitates passage through the cell cycle [51]. The recent conceptualisation of a systems biology mathematical model of mammalian G1 cell cycle progression illustrates a tentative multidisciplinary approach to profiling and customising cell-cycle targeted chemotherapy. This dynamical model integrates current theoretical principles of G1 cell-cycle progression and relies strongly on growth

factor-induced upregulation of cyclin activity to induce oscillation between inactive phosphorylated pRb and active hypophosphorylated pRb [52]. Moreover, the use of antibodies to downregulate cyclin overexpression in cancers may be valuable relative to antibody-mediated enhancement of tumour cell targeting [53]. The concept of selective tumour ablation by application of multiple-effect agents has proved successful in clinical trials with the multi-target tumour growth inhibitor, ZK 304709, which exhibits specificity for CDK-1 and -2 as well as VEGFRs 1, 2, 3 and platelet-derived growth factor receptor. The efficacy of ZK 304709 is consistent with the principle that dual inhibition of cell-cycle progression and neoangiogenesis might produce improved cancer remission through its ability to inhibit vascular permeability, cell cycle molecular targets, as well as tumour growth [54,55]. Thus, multi-target anticancer agents such as ZK 304709 afford a novel approach for disrupting the tumour microenvironment, the site of many oncogenic events and interactions [56]. The caspases are important molecular players in apoptosis signalling and show promise in apoptotic intervention as a basis for targeted cancer treatment [70-73,85-94]. In human cancers, downregulation of caspases blocks apoptosis, thus, potentiating the assertive clonal proliferation of tumour cells. IAPs and survivin increase ubiquitylation of caspases and their degradation by the proteasome [73,95]. IAPs may be used as antiapoptotic decoys because they compete with specific downstream caspase targets that are necessary for apoptosis. IAP antagonists may serve as biological markers to predict therapeutic outcome in cancer patients with high expression levels of IAPs [96,97]. shRNA-knockdown or silencing of p53 and caspases to inhibit apoptosis proved sufficient to delineate apoptosis defects in drug-sensitive and -resistant cancer cells, and may be used to screen for prognostic markers (e.g., caspasedependent cytochrome c-related activation of caspase-3) as well as to design novel apoptogenic (aptoptosis-inducing) or caspase-activation therapeutic strategies [84,104]. Signature gain of oncogenes and loss of tumour suppressor genes indicate loss of UPR functionality in maintaining steady-state levels of normal and mutant proteins, thus, conferring a selective growth advantage on tumour cells and rendering them resistant to apoptosis [17,71,107-111]. Tumour cell resistance to apoptosis is mediated by transcriptional upregulation of antiapoptotic proteins or downregulation of pro-apoptotic proteins through the UPR. The upregulation and activation of NF-κB and downregulation of its inhibitor (I-κB) in many human cancers reflect this concept of apoptosis incompetence [118,121-125]. NF-κB inhibits apoptosis, antagonises p53 [136] and promotes cell division by the transactivation of numerous antiapoptotic target genes such as caspase inhibitors (c-Flip, cIAP-1, cIAP-2 and XIAP) and antiapoptotic members of the Bcl-2 family [113,117,118,126]. Thus, activation of NF-κB has been implicated in poor prognosis. The significance of targeting and downregulating the NF-κB pathway in multiple myeloma and highly incurable


913


mantle cell lymphoma is borne out by the potent effects produced by the combination of inhibitors of IKK1 with melphalan, bortezomib and histone deacetylase inhibitors [128,129]. Moreover, the use of adenoviral expression systems to deliver RNA aptamers that selectively target and inhibit NF-κB activation in human cancers has shown promise, particularly with regard to ablation of drug resistance and angiogenesis induced by NF-κB [130]. The proteasome inhibitor, bortezomib, has been widely studied and numerous clinical trials point to its efficacy in the treatment of multiple myeloma patients, either as a single agent or in combination with standard anticancer agents [1,3,22,142-146,176]. The anticancer activity of proteasome inhibitors generally relate to their ability to promote pro-apoptotic functions and minimise antiapoptotic processes. These include inactivation of NF-κB through decreased degradation of I-κB, reduced NF-κB-dependent upregulation of antiapoptotic regulators such as c-Flip, IAPs, Bcl-2, angiogenic factors (e.g., VEGF), stabilisation of p53, JNK, deregulation of cyclin turnover, downregulation of Mcl-1 and its caspase-dependent degradation, and so on [22,177]. Several significant ERS and UPR role players impact on apoptosis, both in normal physiologic and oncogenic contexts. Figure 1 summarises some of the ERS, UPR and apoptotic role players described in this review that can be targeted for cancer chemotherapy. Thus, components of the UPR in human cancers have proven useful as potential markers for monitoring disease activity, and for predicting and effecting patient survival [8,175,181-183]. Despite the overwhelming accomplishments with bortezomib as a molecularly tailored anticancer agent, many intricacies surround proteasome inhibition, including the complexity of several interconnected cancer signalling pathways that still need to be navigated to identify critical nodes of UPR-mediated apoptosis regulation, which can be targeted to circumvent the progression of apoptosis resistant cancers. 4.

Expert opinion

Drug discovery is a time-honoured theme and process in cancer therapeutics that demands integration of knowledge of the pharmacodynamic interaction between the therapeutic target (tumour milieu) and the drug, robust empirical evaluation of the pharmacokinetics and bioavailability of the drug, lead profiling and optimisation and rational clinical evolution of the drug in terms of efficacy and safety [109]. Together, ERS, UPR and apoptosis signalling cascades (the molecular therapeutic targets) and bortezomib (the drug that attacks the precise molecular pathology of the targets) symbolise a success story in cancer drug discovery, and more especially in the treatment of multiple myeloma. The accumulation of immunoglobulin in multiple myeloma leads to constitutive ERS and upregulation of ERS chaperones and UPR proteins to mediate proteasomal degradation that ensure tumour survival processes. Unabated ERS and constant inhibition of the 914

proteasome by bortezomib define the molecular basis for the sensitivity of myeloma cells to bortezomib. By contrast, solid sarcomas are not as sensitive to bortezomib as myeloma and require an intervention strategy that would sensitise them to bortezomib. A recent study showed that combined treatment of bortezomib-resistant sarcoma cells with the HIV protease inhibitor ritonavir and bortezomib produced synergistic efficacy that mechanistically integrated sustained ERS and UPR signals such as PERK, IRE1 and ATF6, and activated GADD153/CHOP, JNK, caspase-4 and caspase-9 that triggered a high degree of apoptosis [184]. Despite the advances in proteasome inhibition as a molecularly targeted chemotherapeutic modality for multiple myeloma, acquired resistance to bortezomib following continuous exposure and the reported inefficacy of bortezomib in breast cancer call for attention, especially in view of the high clinical attrition rates for new oncology drugs [10,179,185,186]. The development of resistance to proteasome inhibitors may occur through several concerted mechanisms, including enhanced de novo biogenesis of 19S-20S-19S proteasomes, increased expression of the POMP and antiapoptotic Hsp27, and impaired proapoptotic p53 protein upregulation and stabilisation that confer on cancer cells a survival advantage to assert a hyperproliferative and apoptosis-resistant phenotype [180]. It is highly probable that resistance to proteasome inhibition is mediated by a molecular switch, in this case the UPR, that turns on antiapoptotic signals as an adaptive response to unrelenting proteasome stress. Clinical experience with bortezomib in relapsed myeloma patients raised questions about its safety, because side effects such as leukopaenia, infection, herpes zoster, thrombocytopaenia, neuropathy and fatigue were reported [176]. However, the efficacy of bortezomib can be enhanced in synergistic combinations with other anticancer agents such as rituximab, 5-fluorouracil, paclitaxel, doxorubicin, melphalan, intermediate-dose dexamethasone and continuous low-dose oral cyclophosphamide, such that adverse events, including resistance, can be prevented by mandatory antiviral prophylaxis. Therefore, attainment of higher efficacy of bortezomib is possible with combination therapy in regimens with lower drug dosages and with a reduced probability of drug-resistance [22,147,176-178]. Proteasome-based cytoreductive chemotherapy using bortezomib alone or in combination with other drugs exemplifies the efficacy of chemotherapy-induced apoptosis in aggressive and refractory cancers [177,178]. Other milestones in apoptosis-targeted chemotherapy of various cancers are drug design and development towards highly selective multi-target agents, such as cell cycle regulator-antibody-dependent tumour toxicity enhancement [53], concurrent inhibition of cell-cycle progression and neoangiogenesis [54-56,187], combinations of nutlin (an Mdm2-antagonist) with: p53 gene therapy and oncolytic virotherapy of cancer [188]; TRAIL, an inducer of the p53-pathway; gene-specific knockdown of transcription factors with siRNAs [189-191]; and inhibitors of protein–protein


Hiss & Gabriels

• Normal G1/S restriction or progression • Apoptosis sensitive • Normal p53, pRb, Cyclins, CDKs

Normal cell

Mutations DNA damage ERS (Glucose starvation, hypoxia, cellular and ER calcium depletion, high mutation load of unfolded proteins)

• Chemo-and radiation therapy • Genome instability • Apoptosis deregulation • Loss of cell cycle regulation

Gain of function • Cyclins, CDKs, VEGFR, PDGFR, EGFR, IAPs, c-Myc, multiple ER chaperones, Raf-1, Akt, NF-κB, STAT3, c-Flip, HIF-1α, VEGF, Mcl-1, amplified de novo biogenesis of 19S-20S-19S proteasomes and POMP in response to continued proteasome inhibtion, oncoproteins, anti-apoptotic factors, survivin

Tumour cell

Loss of function • INK4 and Cip/Kip, caspases, AIF, Mad1, STAT1, I-κB, NEMO, IKK, PUMA, tumour suppressor proteins, pro-apoptotic factors

Maintenance of the cancer phenotype • Hyperproliferative signature • Apoptotic arrest and resistance • Loss of normal ERS and UPR functionality • Angiogenesis and metatasis • Drug resistance • Poor prognosis

Figure 1. Some of the UPR, ERS and apoptotic role players that can be targeted for cancer chemotherapy. Proteins levels and function in normal cells are tightly regulated by ERS and the UPR. Tumour cells exploit ERS and the UPR to inhibit apoptosis. Transformation from normal to the cancerous state involves complex gain-of-function or loss-of function effects, resulting in a dominant cancer phenotype with a hyperproliferative signature, apoptosis and drug resistance and poor prognosis (for more details, see text in both Parts I and II). ERS: Endoplasmic reticulum stress; UPR: Unfolded protein response.

interactions [192]. These approaches demonstrate the usefulness of systems-based or multi-target drugs to overcome side effects, resistance and poor prognosis that form the responses of complex integrated cancer signalling networks [193]. Moreover, mathematical modelling of cell cycle events to customise cell-cycle, UPR and apoptosis targeted chemotherapy may become a futuristic reality [52]. The next 5 – 10 years may also witness therapeutic drug monitoring and clinical requests for routine ELISA-type or cell-based assays of proteasome levels in blood samples of patients with cancers that overexpress

components of the UPR to predict treatment outcome [175,181,194]. Also, the increasing application of technologies such as siRNA/shRNA-knockdown, aptamers and RNA decoys to selectively silence genes of the major ERS, UPR and apoptotic role players to effect good prognosis will bolster rational drug design strategies [84,150,189-191,195,196]. Central to apoptosis induction and tumour cell killing is the collateral activation and extensive cross-talk among different ERS, UPR and apoptotic signalling cascades. In this regard, co-translocation of caspase-12 and apoptosis-


915


inducing factor to the nucleus as a necessary corollary to ERS (e.g., misfolded proteins, disturbance of ER and cytosolic Ca2+ homeostasis and calpain activation) and reinforcement of UPR-mediated pro-apoptotic signalling may be of note [197]. We foresee exciting future prospects for specific cancer drug targeting of the ERS, UPR and apoptotic pathways. Bibliography 1.

2.

3.

4.

5.

6.

Nowis D, McConnell EJ, Dierlam L, et al. TNF potentiates anticancer activity of bortezomib (Velcade) through reduced expression of proteasome subunits and dysregulation of unfolded protein response. Int J Cancer 2007;121(2):431-41 Milano A, Iaffaioli RV, Caponigro F. The proteasome: a worthwhile target for the treatment of solid tumours? Eur J Cancer 2007;43(7):1125-33 Neukirchen J, Meier A, Rohrbeck A, et al. The proteasome inhibitor bortezomib acts differently in combination with p53 gene transfer or cytotoxic chemotherapy on NSCLC cells. Cancer Gene Ther 2007;14(4):431-9 Goldberg AL. Functions of the proteasome: from protein degradation and immune surveillance to cancer therapy. Biochem Soc Trans 2007;35(Pt 1):12-7 Sterz J, Metzler IV, Hahne J-C, et al. The potential of proteasome inhibitors in cancer therapy. Exp Opin Invest Drugs 2008;17(6):879-95

Declaration of interest The authors state no conflict of interest and have received no payment in preparation of this manuscript. This work was supported in part by funds from the University of the Western Cape, the University of Cape Town and the Ackerman Family Educational Trust.

12.

Thorpe JA, Christian PA, Schwarze SR. Proteasome inhibition blocks caspase-8 degradation and sensitizes prostate cancer cells to death receptor-mediated apoptosis. Prostate 2008;68(2):200-9

24.

Farras R, Bossis G, Andermarcher E, et al. Mechanisms of delivery of ubiquitylated proteins to the proteasome: new target for anti-cancer therapy? Crit Rev Oncol Hematol 2005;54(1):31-51

13.

Hallett WH, Ames E, Motarjemi M, et al. Sensitization of tumor cells to NK cell-mediated killing by proteasome inhibition. J Immunol 2008;180(1):163-70

25.

Li B, Jia N, Waning DL, et al. Cul4A is required for hematopoietic stem-cell engraftment and self-renewal. Blood 2007;110(7):2704-7

14.

Rosa DD, Ismael G, Lago LD, Awada A. Molecular-targeted therapies: lessons from years of clinical development. Cancer Treat Rev 2008;34(1):61-80

26.

Waning DL, Li B, Jia N, et al. Cul4A is required for hematopoietic cell viability and its deficiency leads to apoptosis. Blood 2008;112(2):320-9

15.

Hampton RY. ER stress response: getting the UPR hand on misfolded proteins. Curr Biol 2000;10:R518-R21

27.

16.

Nalepa G, Harper W. Therapeutic anti-cancer targets upstream of the proteasome. Cancer Treat Rev 2003;29:49-57

Heuzé ML, Lamsoul I, Moog-Lutz C, Lutz PG. Ubiquitin-mediated proteasomal degradation in normal and malignant hematopoiesis. Blood Cells Mol Dis 2008;40(2):200-10

28.

Brewer JW, Diehl JA. PERK mediates cell-cycle exit during the mammalian unfolded protein response. Proc Natl Acad Sci USA 2000;97(23):12625-30

29.

Brewer JW, Hendershot LM, Sherr CJ, Diehl JA. Mammalian unfolded protein response inhibits cyclin D1 translation and cell-cycle progression. Proc Natl Acad Sci USA 1999;96(15):8505-10

30.

Zhang F, Hamanaka RB, Bobrovnikova-Marjon E, et al. Ribosomal stress couples the unfolded protein response to p53-dependent cell cycle arrest. J Biol Chem 2006;281(40):30036-45

31.

Hamanaka RB, Bennett BS, Cullinan SB, Diehl JA. PERK and GCN2 contribute to eIF2alpha phosphorylation and cell cycle arrest after activation of the unfolded protein response pathway. Mol Biol Cell 2005;16(12):5493-501

32.

Bicknell AA, Babour A, Federovitch CM, Niwa M. A novel role in cytokinesis reveals a housekeeping function for the unfolded protein response. J Cell Biol 2007;177(6):1017-27

33.

Aggarwal BB, Banerjee S, Bharadwaj U, et al. Curcumin induces the degradation of cyclin E expression through ubiquitin-dependent pathway and

17.

18.

Crawford LJ, Walker B, Irvine AE. Proteasome inhibitors: a therapeutic strategy for haematological malignancy. Front Biosci 2008;13:4285-96

Nalepa G, Rolfe M, Harper JW. Drug discovery in the ubiquitin–proteasome system. Nat Rev Drug Discov 2006;5:596-613 Zavrski I, Jakob C, Kaiser M, et al. Molecular and clinical aspects of proteasome inhibition in the treatment of cancer. Recent Results Cancer Res 2007;176:165-76

7.

Daviet L, Colland F. Targeting ubiquitin specific proteases for drug discovery. Biochimie 2008;90(2):270-83

19.

Vink J, Cloos J, Kaspers GJL. Proteasome inhibition as novel treatment strategy in leukaemia. Br J Haematol 2006;134:253-62

8.

Burger AM. Highlights in experimental therapeutics. Cancer Lett 2007;245(1-2):11-21

20.

Voorhees PM, Orlowski RZ. The proteasome and proteasome inhibitors in cancer therapy. Annu Rev Pharmacol Toxicol 2006;46(1):189-213

9.

10.

11.

916

Strasser A, Puthalakath H. Fold up or perish: unfolded protein response and chemotherapy. Cell Death Differ 2008;15(2):223-5 Smith L, Lind MJ, Drew PJ, Cawkwell L. The putative roles of the ubiquitin/ proteasome pathway in resistance to anticancer therapy. Eur J Cancer 2007;43(16):2330-8 Moran E, Nencioni A. The role of proteasome in malignant diseases. J Buon 2007;12(Suppl 1):S95-S99

21.

Kuhn DJ, Zeger EL, Orlowski RZ. Proteasome inhibitors and modulators of heat shock protein function. Update Cancer Ther 2006;1(2):91-116

22.

Nencioni A, Grünebach F, Patrone F, et al. Proteasome inhibitors: Antitumor effects and beyond. Leukemia 2007;21:30-6

23.

Ovaa H. Active-site directed probes to report enzymatic action in the ubiquitin proteasome system. Nat Rev Cancer 2007;7(8):613-20


Hiss & Gabriels

up-regulates cyclin-dependent kinase inhibitors p21 and p27 in multiple human tumor cell lines. Biochem Pharmacol 2007;73(7):1024-32 34.

35.

36.

37.

38.

39.

40.

41.

Akatsu Y, Saikawa Y, Kubota T, et al. Predictive value of GADD153, p21 and c-Jun for chemotherapy response in gastric cancer. Cancer Sci 2007;98(5):707-15 Avkin S, Sevilya Z, Toube L, et al. p53 and p21 regulate error-prone DNA repair to yield a lower mutation load. Mol Cell 2006;22(3):407-13 Bai M, Tsanou E, Skyrlas A, et al. Alterations of the p53, Rb and p27 tumor suppressor pathways in diffuse large B-cell lymphomas. Anticancer Res 2007;27(4B):2345-52 Cao W, Chi W-H, Wang J, et al. TNF-[alpha] promotes doxorubicin-induced cell apoptosis and anti-cancer effect through downregulation of p21 in p53-deficient tumor cells. Biochem Biophys Res Commun 2005;330(4):1034-40

for cancer therapy. Drug Resist Updat 2007;10(1-2):13-29 44.

Mathew R, Arora S, Khanna R, et al. Alterations in p53 and pRb pathways and their prognostic significance in oesophageal cancer. Eur J Cancer 2002;38(6):832-41

45.

Matta H, Chaudhary PM. The proteasome inhibitor bortezomib (PS-341) inhibits growth and induces apoptosis in primary effusion lymphoma cells. Cancer Biol Ther 2005;4(1):77-82

Lin Y-C, Wang F-F. Mechanisms underlying the pro-survival pathway of p53 in suppressing mitotic death induced by adriamycin. Cell Signal 2008;20(1):258-67

43.

Maddika S, Ande SR, Panigrahi S, et al. Cell survival, cell death and cell cycle pathways are interconnected: Implications

Akli S, Keyomarsi K. Cyclin E and its low molecular weight forms in human cancer and as targets for cancer therapy. Cancer Biol Ther 2003;2(4(Suppl 1)):S38-47

59.

Strasser S, Maier S, Leisser C, et al. 5-FdUrd–araC heterodinucleoside re-establishes sensitivity in 5-FdUrd and AraC-resistant MCF-7 breast cancer cells overexpressing ErbB2. Differentiation 2006;74:488-98

47.

Rakitina TV, Vasilevskaya IA, O’Dwyer PJ. Inhibition of G1/S transition potentiates oxaliplatin-induced cell death in colon cancer cell lines. Biochem Pharmacol 2007;73(11):1715-26

60.

Dai L, Wang X, Yao X, et al. Antisense oligonucleotide targeting p53 increased apoptosis of MCF-7 cells induced by ionizing radiation. Acta Pharmacol Sin 2006;27(11):1453-8

Grant S, Roberts J. The use of cyclin-dependent kinase inhibitors alone or in combination with established cytotoxic drugs in cancer chemotherapy. Drug Resist Updat 2003;6:15-26

61.

Nakayama K, Nakayama K. Ubiquitin ligases: cell-cycle control and cancer. Nat Rev Cancer 2006;6:369-81

Shapiro G. Preclinical and clinical development of the cyclindependent kinase inhibitor flavopiridol. Clin Cancer Res 2004;10:4270s-5s

62.

Motwani M, Rizzo C, Sirotnak F, et al. Flavopiridol enhances the effect of docetaxel in vitro and in vivo in human gastric cancer cells. Mol Cancer Ther 2003;2:549-55

63.

Osoegawa A, Yoshino I, Tanaka S, et al. Regulation of p27 by S-phase kinase-associated protein 2 is associated with aggressiveness in non-small-cell lung cancer. J Clin Oncol 2004;22:4165-73

64.

Ben-Izhak O, Lahav-Baratz S, Meretyk S, et al. Inverse relationship between Skp2 ubiquitin ligase and the cyclin dependent kinase inhibitor p27Kip1 in prostate cancer. J Urol 2003;170:241-5

65.

Nickeleit I, Zender S, Sasse F, et al. Argyrin A reveals a critical role for the tumor suppressor protein p27kip1 in mediating antitumor activities in response to proteasome inhibition. Cancer Cell 2008;14(1):23-35

66.

Flemming A. Anticancer Drugs: proteasome inhibitor unleashes three-pronged attack. Nat Rev Drug Discov 2008;7(9):730-1

67.

McConkey DJ. A novel role for a familiar protein in apoptosis induced by proteasome inhibition. Cancer Cell 2008;14(1):1-2

68.

Devoy A, Soane T, Welchman R, Mayer RJ. The ubiquitin-proteasome system and cancer. Essays Biochem 2005;41:187-203

48.

Ding H, Duan W, Zhu W-G, et al. p21 response to DNA damage induced by genistein and etoposide in human lung cancer cells. Biochem Biophys Res Commun 2003;305(4):950-6

42.

58.

Nam EJ, Kim YT. Alteration of cell-cycle regulation in epithelial ovarian cancer. Int J Gynecol Cancer 2008;18(6):1169-82

49.

Kolomeichuk SN, Bene A, Upreti M, et al. Induction of apoptosis by vinblastine via c-Jun autoamplification and p53-independent down-regulation of p21WAF1/CIP1. Mol Pharmacol 2008;73(1):128-36

Matsumoto M, Furihata M, Ishikawa T. Comparison of deregulated expression of cyclin D1 and cyclin E with that of cyclin-dependent kinase 4 (CDK4) and CDK2 in human oesophageal squamous cell carcinoma. Br J Cancer 1999;80:256

46.

Chen C-Y, Hsu Y-L, Chen Y-Y, et al. Isokotomolide A, a new butanolide extracted from the leaves of Cinnamomum kotoense, arrests cell cycle progression and induces apoptosis through the induction of p53/p21 and the initiation of mitochondrial system in human non-small cell lung cancer A549 cells. Eur J Pharmacol 2007;574(2-3):94-102

Huang TG, Ip SM, Yeung WSB, Ngan HYS. Changes in p21WAF1, pRb, Mdm-2, Bax and Bcl-2 expression in cervical cancer cell lines transfected with a p53 expressing adenovirus. Eur J Cancer 2000;36(2):249-56

57.

50.

Menon S, Goswami P. A redox cycle within the cell cycle: ring in the old with the new. Oncogene 2007;26:1101-9

51.

Weinberg R. The retinoblastoma protein and cell cycle control. Cell 1995;81:323

52.

Haberichter T, Madge B, Christopher RA, et al. A systems biology dynamical model of mammalian G1 cell cycle progression. Mol Syst Biol 2007;3:1-8

53.

Sanz L, Cuesta ÁM, Compte M, Álvarez-Vallina L. Antibody engineering: facing new challenges in cancer therapy. Acta Pharmacol Sin 2005;26(6):641-8

54.

Broxterman HJ, Georgopapadakou NH. Anticancer therapeutics: “Addictive” targets, multi-targeted drugs, new drug combinations. Drug Resist Update 2005;8:183-97

55.

Siemeister G, Briem H, Brumby T. ZK 304709: the oral multi-target tumor growth inhibitor™ (MTGI™) acts via inhibition of cell cycle progression and tumor-induced neoangiogenesis [abstract # 5842]. Am Assoc Cancer Res 2005;46

56.

Bogenrieder T, Herlyn M. Axis of evil: molecular mechanisms of cancer metastasis. Oncogene 2003;22:6524-36


917


69.

Boelens J, Lust S, Offner F, et al. Review. The endoplasmic reticulum: a target for new anticancer drugs. In Vivo 2007;21(2):215-26

70.

Lamkanfi M, Festjens N, Declercq W, et al. Caspases in cell survival, proliferation and differentiation. Cell Death Differ 2007;14:44-55

71.

Jin Z, El-Deiry WS. Overview of cell death signaling pathways. Cancer Biol Ther 2005;4(2):e50-74

72.

Taylor RC, Cullen SP, Martin SJ. Apoptosis: controlled demolition at the cellular level. Nat Rev Mol Cell Biol 2008;9(3):231-41

73.

Kumar S. Caspase function in programmed cell death. Cell Death Differ 2007;14:32-43

74.

Guerrero AD, Chen M, Wang J. Delineation of the caspase-9 signaling cascade. Apoptosis 2008;13(1):177-86

75.

Hetfeld BK, Peth A, Sun XM, et al. The COP9 signalosome-mediated deneddylation is stimulated by caspases during apoptosis. Apoptosis 2008;13(2):187-95

76.

Schroder M, Kaufman RJ. ER stress and the unfolded protein response. Mutat Res 2005;569(1-2):29-63

77.

Biagioli M, Pifferi S, Ragghianti M, et al. Endoplasmic reticulum stress and alteration in calcium homeostasis are involved in cadmium-induced apoptosis. Cell Calcium 2008;43(2):184-95

78.

79.

80.

81.

918

82.

83.

Deniaud A, Sharaf el dein O, Maillier E, et al. Endoplasmic reticulum stress induces calcium-dependent permeability transition, mitochondrial outer membrane permeabilization and apoptosis. Oncogene 2008;27(3):285-99

84.

Ehrhardt H, Hacker S, Wittmann S, et al. Cytotoxic drug-induced, p53-mediated upregulation of caspase-8 in tumor cells. Oncogene 2008;27(6):783-93

85.

Lien YC, Kung HN, Lu KS, et al. Involvement of endoplasmic reticulum stress and activation of MAP kinases in beta-lapachone-induced human prostate cancer cell apoptosis. Histol Histopathol 2008;23(11):1299-308 Rao RV, Poksay KS, Castro-Obregon S, et al. Molecular components of a cell death pathway activated by endoplasmic reticulum stress. J Biol Chem 2004;279(1):177-87

Abdelrahim M, Newman K, Vanderlaag K, et al. 3,3′-diindolylmethane (DIM) and its derivatives induce apoptosis in pancreatic cancer cells through endoplasmic reticulum stress-dependent upregulation of DR5. Carcinogenesis 2006;27(4):717-28

86.

Alkhalaf M, El-Mowafy A, Renno W, et al. Resveratrol-induced apoptosis in human breast cancer cells is mediated primarily through the caspase-3-dependent pathway. Arch Med Res 2008;39(2):162-8

87.

Barbour KW, Berger FG. Cell death in response to antimetabolites directed at thymidylate synthase. Cancer Chemother Pharmacol 2008;61(2):189-201

88.

Chiang PC, Chien CL, Pan SL, et al. Induction of endoplasmic reticulum stress and apoptosis by a marine prostanoid in human hepatocellular carcinoma. J Hepatol 2005;43(4):679-86 Yeh TC, Chiang PC, Li TK, et al. Genistein induces apoptosis in human hepatocellular carcinomas via interaction of endoplasmic reticulum stress and mitochondrial insult. Biochem Pharmacol 2007;73(6):782-92

Rao RV, Niazi K, Mollahan P, et al. Coupling endoplasmic reticulum stress to the cell-death program: A novel HSP90-independent role for the small chaperone protein p23. Cell Death Differ 2006;13(3):415-25

89.

90.

91.

92.

Anding AL, Chapman JS, Barnett DW, et al. The unhydrolyzable fenretinide analogue 4-hydroxybenzylretinone induces the proapoptotic genes GADD153 (CHOP) and Bcl-2-binding component 3 (PUMA) and apoptosis that is caspase- dependent and independent of the retinoic acid receptor. Cancer Res 2007;67(13):6270-7 Bokelmann I, Mahlknecht U. Valproic acid sensitizes chronic lymphocytic leukemia cells to apoptosis and restores the balance between pro- and antiapoptotic proteins. Mol Med 2008;14(1-2):20-7 Can I, Tahara-Hanaoka S, Hitomi K, et al. Caspase-independent cell death by CD300LF (MAIR-V), an inhibitory immunoglobulin-like receptor on myeloid cells. J Immunol 2008;180(1):207-13 Galbán S, Brady GF, Duckett CS. Caspases and IAPs: a dance of death ensures cell survival. Mol Cell 2008;32(4):462-3 Grozio A, Paleari L, Catassi A, et al. Natural agents targeting the alpha7-nicotinic-receptor in NSCLC:


A promising prospective in anti-cancer drug development. Int J Cancer 2008;122(8):1911-5 93.

Klampfer L. The role of signal transducers and activators of transcription in colon cancer. Front Biosci 2008;13:2888-99

94.

Ondrouskova E, Soucek K, Horvath V, Smarda J. Alternative pathways of programmed cell death are activated in cells with defective caspase-dependent apoptosis. Leuk Res 2008;32(4):599-609

95.

Callus B, Vaux D. Caspase inhibitors: viral, cellular and chemical. Cell Death Differ 2007;14:73-8

96.

Reed JC. Drug Insight: cancer therapy strategies based on restoration of endogenous cell death mechanisms. Nat Clin Pract Oncol 2006;3(7):388-98

97.

Xu L, Zhu J, Hu X, et al. c-IAP1 cooperates with Myc by acting as a ubiquitin ligase for Mad1. Mol Cell 2007;28(5):914-22

98.

Hjerpe R, Rodríguez MS. Alternative UPS drug targets upstream the 26S proteasome. Int J Biochem Cell Biol 2008;40(6-7):1126-40

99.

Mayer RJ, Fujita J. Gankyrin, the 26 S proteasome, the cell cycle and cancer. Biochem Soc Trans 2006;34(5):746-8

100. Cillessen SA, Reed JC, Welsh K, et al. Small-molecule XIAP antagonist restores caspase-9 mediated apoptosis in XIAP-positive diffuse large B-cell lymphoma cells. Blood 2008;111(1):369-75 101. Dasmahapatra G, Lembersky D, Rahmani M, et al. Bcl-2 antagonists interact synergistically with bortezomib in DLBCL cells in association with JNK activation and induction of ER stress. Cancer Biol Ther 2009;8(9):820-2 102. Caraglia M, Marra M, Budillon A, et al. Chemotherapy regimen GOLF induces apoptosis in colon cancer cells through multi-chaperone complex inactivation and increased Raf-1 ubiquitin-dependent degradation. Cancer Biol Ther 2005;4(10):1159-67 103. Hoffarth S, Zitzer A, Wiewrodt R, et al. pp32/PHAPI determines the apoptosis response of non-small-cell lung cancer. Cell Death Differ 2008;15(1):161-70 104. Meyer LH, Queudeville M, Eckhoff SM, et al. Intact apoptosis signaling in myeloid leukemia cells determines treatment

Hiss & Gabriels

outcome in childhood AML. Blood 2008;111(5):2899-903 105. Kang DW, Choi CH, Park JY, et al. Ciglitazone induces caspase-independent apoptosis through down-regulation of XIAP and survivin in human glioma cells. Neurochem Res 2008;33(3):551-61 106. McLean L, Soto U, Agama K, et al. Aminoflavone induces oxidative DNA damage and reactive oxidative species-mediated apoptosis in breast cancer cells. Int J Cancer 2008;122(7):1665-74 107. Song H, Hollstein M, Xu Y. p53 gain-of-function cancer mutants induce genetic instability by inactivating ATM. Nat Cell Biol 2007;9(5):573-80 108. Fuster JJ, Sanz-González SM, Moll UM, Andrés V. Classic and novel roles of p53: Prospects for anticancer therapy. Trends Mol Med 2007;13(5):192-9 109. Collins I, Workman P. New approaches to molecular cancer therapeutics. Nat Chem Biol 2006;2(12):689-700 110. Eastman A, Perez RP. New targets and challenges in the molecular therapeutics of cancer. Br J Clin Pharmacol 2006;62(1):5-14 111. Zinszner H, Kuroda M, Wang X, et al. CHOP is implicated in programmed cell death in response to impaired function of the endoplasmic reticulum. Genes Dev 1998;12:982-95 112. Xu C, Bailly-Maitre B, Reed JC. Endoplasmic reticulum stress: cell life and death decisions. J Clin Invest 2005;115(10):2656-64 113. Tabruyn SP, Griffioen AW. A new role for NF-κB in angiogenesis inhibition. Cell Death Differ 2007;14:1393-7 114. Karin M, Greten FR. NF-κB: linking inflammation and immunity to cancer development and progression. Nat Rev Immunol 2005;5:749-59 115. Leclerc D, Rozen R. Endoplasmic reticulum stress increases the expression of methylenetetrahydrofolate reductase through the IRE1 transducer. J Biol Chem 2008;283(6):3151-60 116. Kaneko M, Niinuma Y, Nomura Y. Activation signal of nuclear factor-kappa B in response to endoplasmic reticulum stress is transduced via IRE1 and tumor necrosis factor receptor-associated factor 2. Biol Pharm Bull 2003;26(7):931-5 117. Yoshimura A. Signal transduction of inflammatory cytokines and tumor

development. Cancer Sci 2006;97(6):439-47 118. Fribley A, Wang C-Y. Proteasome inhibitor induces apoptosis through induction of endoplasmic reticulum stress. Cancer Biol Ther 2006;5(7):745-8 119. Heyninck K, Beyaert R. A20 inhibits NF-kappaB activation by dual ubiquitin-editing functions. Trends Biochem Sci 2005;30(1):1-4 120. Zhang HG, Wang J, Yang X, et al. Regulation of apoptosis proteins in cancer cells by ubiquitin. Oncogene 2004;23(11):2009-15 121. Wang C, Mayo M, Baldwin A. TNF- and cancer therapy-induced apoptosis: Potentiation by inhibition of NF-kappaB. Science 1996;274:784-7 122. Wang C, Cusack J, Liu R, Baldwin A. Control of inducible chemoresistance: Enhanced anti-tumor therapy via increased apoptosis through inhibition of NF-kB. Nat Med 1999;5:412-7 123. Wang C, Guttridge D, Mayo M, Baldwin A. NF-kappaB induces expression of the Bcl-2 homologue A1/Bfl-1 to preferentially suppress chemotherapy-induced apoptosis. Mol Cell Biol 1999;19:5923-9 124. Hideshima T, Richardson P, Chauhan D, et al. The proteasome inhibitor PS-341 inhibits growth, induces apoptosis, and overcomes drug resistance in human multiple myeloma cells. Cancer Res 2001;61:3071-6 125. Ondrey F, Dong G, Sunwoo J, et al. Constitutive activation of transcription factors NF-(kappa)B, AP-1, and NF-IL6 in human head and neck squamous cell carcinoma cell lines that express pro-inflammatory and pro-angiogenic cytokines. Mol Carcinog 1999;26:119-29 126. Nencioni A, Grunebach F, Patrone F, et al. Proteasome inhibitors: Antitumor effects and beyond. Leukemia 2007;21(1):30-6 127. Fahy BN, Schlieman MG, Mortenson MM, et al. Targeting BCL-2 overexpression in various human malignancies through NF-kappaB inhibition by the proteasome inhibitor bortezomib. Cancer Chemother Pharmacol 2005;56(1):46-54 128. Jourdan M, Moreaux J, Vos JD, et al. Targeting NF-kB pathway with an IKK2 inhibitor induces inhibition of multiple myeloma cell growth. Br J Haematol 2007;138:160-8


129. Heider U, von Metzler I, Kaiser M, et al. Synergistic interaction of the histone deacetylase inhibitor SAHA with the proteasome inhibitor bortezomib in mantle cell lymphoma. Eur J Haematol 2008;80(2):133-42 130. Mi J, Zhang X, Rabbani ZN, et al. RNA aptamer-targeted inhibition of NF-kappaB suppresses non-small cell lung cancer resistance to doxorubicin. Mol Ther 2008;16(1):66-73 131. Frelin C, Imbert V, Bottero V, et al. Inhibition of the NF-kappaB survival pathway via caspase-dependent cleavage of the IKK complex scaffold protein and NF-kappaB essential modulator NEMO. Cell Death Differ 2008;15(1):152-60 132. Olivier S, Robe P, Bours V. Can NF-kappaB be a target for novel and efficient anti-cancer agents? Biochem Pharmacol 2006;72(9):1054-68 133. Ostrowska H. The ubiquitin-proteasome system: a novel target for anticancer and anti-inflammatory drug research. Cell Mol Biol Lett 2008;13(3):353-65 134. Zavrski I, Kleeberg L, Kaiser M, et al. Proteasome as an emerging therapeutic target in cancer. Curr Pharm Des 2007;13(5):471-85 135. Zavrski I, Jakob C, Schmid P, et al. Proteasome: an emerging target for cancer therapy. Anticancer Drugs 2005;16(5):475-81 136. Taneja P, Mallakin A, Matise LA, et al. Repression of Dmp1 and Arf transcription by anthracyclins: critical roles of the NF-kappaB subunit p65. Oncogene 2007;26(53):7457-66 137. Concannon C, Koehler B, Reimertz C, et al. Apoptosis induced by proteasome inhibition in cancer cells: Predominant role of the p53/PUMA pathway. Oncogene 2007;26:1681-92 138. Nakamura T, Tanaka K, Matsunobu T, et al. The mechanism of cross-resistance to proteasome inhibitor bortezomib and overcoming resistance in Ewing’s family tumor cells. Int J Oncol 2007;31(4):803-11 139. Ishii Y, Waxman S, Germain D. Targeting the ubiquitin-proteasome pathway in cancer therapy. Anticancer Agents Med Chem 2007;7(3):359-65 140. Meng L, Mohan R, Kwok B, et al. Epoxomicin, a potent and selective proteasome inhibitor, exhibits in vivo antiinflammatory activity. Proc Natl Acad Sci USA 1999;96:10403-8

919


141. Lass A, McConnell E, Nowis D, et al. A novel function of VCP (valosin-containing protein; p97) in the control of N-glycosylation of proteins in the endoplasmic reticulum. Arch Biochem Biophys 2007;462(1):62-73 142. Richardson PG. A phase 2 study of bortezomib in relapsed, refractory myeloma. N Engl J Med 2003;348:2609-17 143. Richardson PG. Bortezomib or high-dose dexamethasone for relapsed multiple myeloma. N Engl J Med 2005;352:2487-98 144. Richardson PG, Mitsiades C. Bortezomib: proteasome inhibition as an effective anticancer therapy. Future Oncol 2005;1(2):161-71 145. Richardson PG, Mitsiades C, Hideshima T, Anderson KC. Bortezomib: proteasome inhibition as an effective anticancer therapy. Annu Rev Med 2006;57:33-47 146. Richardson PG, Mitsiades C, Schlossman R, et al. New drugs for myeloma. Oncologist 2007;12(6):664-89 147. Fujita T, Doihara H, Washio K, et al. Antitumor effects and drug interactions of the proteasome inhibitor bortezomib (PS341) in gastric cancer cells. Anticancer Drugs 2007;18(6):677-86 148. Boccadoro M, Morgan G, Cavenagh J. Preclinical evaluation of the proteasome inhibitor bortezomib in cancer therapy. Cancer Cell Int 2005;5(1):18 149. Roccaro AM, Hideshima T, Richardson PG, et al. Bortezomib as an antitumor agent. Curr Pharm Biotechnol 2006;7(6):441-8 150. Kardosh A, Golden EB, Pyrko P, et al. Aggravated endoplasmic reticulum stress as a basis for enhanced glioblastoma cell killing by bortezomib in combination with celecoxib or its non-coxib analogue, 2,5-dimethyl-celecoxib. Cancer Res 2008;68(3):843-51 151. Nikrad M, Johnson T, Puthalalath H, et al. The proteasome inhibitor bortezomib sensitizes cells to killing by death receptor ligand TRAIL via BH3-only proteins Bik and Bim. Mol Cancer Ther 2005;4(3):443-9 152. Fennell DA, Chacko A, Mutti L. BCL-2 family regulation by the 20S proteasome inhibitor bortezomib. Oncogene 2008;27(9):1189-97 153. Fribley AM, Evenchik B, Zeng Q, et al. Proteasome inhibitor PS-341 induces

920

apoptosis in cisplatin-resistant squamous cell carcinoma cells by induction of Noxa. J Biol Chem 2006;281(42):31440-7 154. Li J, Lee B, Lee AS. Endoplasmic reticulum stress-induced apoptosis: multiple pathways and activation of p53-up-regulated modulator of apoptosis (PUMA) and NOXA by p53. J Biol Chem 2006;281(11):7260-70 155. Nikiforov MA, Riblett M, Tang WH, et al. Tumor cell-selective regulation of NOXA by c-MYC in response to proteasome inhibition. Proc Natl Acad Sci USA 2007;104(49):19488-93 156. Chauhan D, Hideshima T, Anderson K. Proteasome inhibition in multiple myeloma: therapeutic implication. Annu Rev Pharmacol Toxicol 2005;45:465-76 157. Chauhan D, Singh A, Brahmandam M, et al. Combination of proteasome inhibitors bortezomib and NPI-0052 trigger in vivo synergistic cytotoxicity in multiple myeloma. Blood 2008;111(3):1654-64 158. Miasari M, Puthalakath H, Silke J. Ubiquitylation and cancer development. Curr Cancer Drug Targets 2008;8(2):118-23 159. Chen C, Kouroukis CT, White D, et al. Bortezomib in relapsed or refractory Waldenstrom’s cacroglobulinemia. Clin Lymphoma Myeloma 2009;9(1):74-6 160. Frankel SR. Oblimersen sodium (G3139 Bcl-2 antisense oligonucleotide) therapy in Waldenstrom’s macroglobulinemia: a targeted approach to enhance apoptosis. Semin Oncol 2003;30(2):300-4 161. Gertz MA, Geyer SM, Badros A, et al. Early results of a phase I trial of oblimersen sodium for relapsed or refractory Waldenstrom’s macroglobulinemia. Clin Lymphoma 2005;5(4):282-4 162. Mitsiades CS, Mitsiades N, Richardson PG, et al. Novel biologically based therapies for Waldenstrom’s macroglobulinemia. Semin Oncol 2003;30(2):309-12 163. Roccaro AM, Leleu X, Sacco A, et al. Resveratrol exerts antiproliferative activity and induces apoptosis in Waldenstrom’s macroglobulinemia. Clin Cancer Res 2008;14(6):1849-58 164. Roccaro AM, Vacca A, Ribatti D. Bortezomib in the treatment of cancer. Recent Patents Anticancer Drug Discov 2006;1(3):397-403 Expert Opin. Drug Discov. (2009) 4(9)

165. Zeldis JB, Schafer PH, Bennett BL, et al. Potential new therapeutics for Waldenstrom’s macroglobulinemia. Semin Oncol 2003;30(2):275-81 166. Hatjiharissi E, Ngo H, Leontovich AA, et al. Proteomic analysis of waldenstrom macroglobulinemia. Cancer Res 2007;67(8):3777-84 167. Leleu X, Xu L, Jia X, et al. Endoplasmic reticulum stress is a target for therapy in Waldenstrom macroglobulinemia. Blood 2009;113(3):626-34 168. Aloy MT, Hadchity E, Bionda C, et al. Protective role of Hsp27 protein against gamma radiation-induced apoptosis and radiosensitization effects of Hsp27 gene silencing in different human tumor cells. Int J Radiat Oncol Biol Phys 2008;70(2):543-53 169. Yaglom JA, Gabai VL, Sherman MY. High levels of heat shock protein Hsp72 in cancer cells suppress default senescence pathways. Cancer Res 2007;67(5):2373-81 170. O’Callaghan-Sunol C, Gabai VL, Sherman MY. Hsp27 modulates p53 signaling and suppresses cellular senescence. Cancer Res 2007;67(24):11779-88 171. Schmitt E, Gehrmann M, Brunet M, et al. Intracellular and extracellular functions of heat shock proteins: Repercussions in cancer therapy. J Leukoc Biol 2007;81(1):15-27 172. Hiss DC, Gabriels GA, Folb PI. Combination of tunicamycin with anticancer drugs synergistically enhances their toxicity in multidrug-resistant human ovarian cystadenocarcinoma cells. Cancer Cell Int 2007;7(1):5 173. Loo T, Clarke D. The human multidrugresistance P-glycoprotein is inactive when its maturation is inhibited: potential for a role in cancer chemotherapy. FASEB J 1999;13(13):1724-32 174. Zhang Z, Wu J-Y, Hait WN, Yang J-M. Regulation of the stability of P-glycoprotein by ubiquitination. Mol Pharmacol 2004;66(3):395-403 175. Wawrzynczak E. Prognostic proteasome. Nat Rev Cancer 2007;7(1):6-7 176. Kropff M, Bisping G, Schuck E, et al. Bortezomib in combination with intermediate-dose dexamethasone and continuous low-dose oral cyclophosphamide for relapsed multiple myeloma. Br J Haematol 2007;138(3):330-7

Hiss & Gabriels

177. Podar K, Gouill SL, Zhang J, et al. A pivotal role for Mcl-1 in Bortezomib-induced apoptosis. Oncogene 2008;27(6):721-31 178. Wang M, Han XH, Zhang L, et al. Bortezomib is synergistic with rituximab and cyclophosphamide in inducing apoptosis of mantle cell lymphoma cells in vitro and in vivo. Leukemia 2008;22(1):179-85 179. Xu H, Ju D, Jarois T, Xie Y. Diminished feedback regulation of proteasome expression and resistance to proteasome inhibitors in breast cancer cells. Breast Cancer Res Treat 2008;107(2):267-74 180. Fuchs D, Berges C, Opelz G, et al. Increased expression and altered subunit composition of proteasomes induced by continuous proteasome inhibition establish apoptosis resistance and hyperproliferation of Burkitt lymphoma cells. J Cell Biochem 2008;103(1):270-83

bortezomib-induced apoptosis. Mol Cancer Ther 2008;7(7):1940-8 185. Yang CH, Gonzalez-Angulo AM, Reuben JM, et al. Bortezomib (VELCADE®) in metastatic breast cancer: Pharmacodynamics, biological effects, and prediction of clinical benefits. Ann Oncol 2006;17(5):813-7 186. Kola I, Landis J. Can the pharmaceutical industry reduce attrition rates? Nat Rev Drug Discov 2004;3:711-5 187. Secchiero P, Corallini F, Gonelli A, et al. Antiangiogenic activity of the MDM2 antagonist nutlin-3. Circ Res 2007;100(1):61-9 188. Graat HC, Carette JE, Schagen FH, et al. Enhanced tumor cell kill by combined treatment with a small-molecule antagonist of mouse double minute 2 and adenoviruses encoding p53. Mol Cancer Ther 2007;6(5):1552-61

181. Jakob C, Egerer K, Liebisch P, et al. Circulating proteasome levels are an independent prognostic factor for survival in multiple myeloma. Blood 2007;109(5):2100-5

189. Secchiero P, Zerbinati C, di Iasio MG, et al. Synergistic cytotoxic activity of recombinant TRAIL plus the non-genotoxic activator of the p53 pathway nutlin-3 in acute myeloid leukemia cells. Curr Drug Metab 2007;8(4):395-403

182. Golab J, Bauer TM, Daniel V, Naujokat C. Role of the ubiquitin-proteasome pathway in the diagnosis of human diseases. Clin Chim Acta 2004;340(1-2):27-40

190. Secchiero P, Zerbinati C, Melloni E, et al. The MDM-2 antagonist nutlin-3 promotes the maturation of acute myeloid leukemic blasts. Neoplasia 2007;9(10):853-61

183. Yang Y, Kitagaki J, Wang H, et al. Targeting the ubiquitin-proteasome system for cancer therapy. Cancer Sci 2009;100(1):24-8

191. Stuhmer T, Chatterjee M, Hildebrandt M, et al. Nongenotoxic activation of the p53 pathway as a therapeutic strategy for multiple myeloma. Blood 2005;106(10):3609-17

184. Kraus M, Malenke E, Gogel J, et al. Ritonavir induces endoplasmic reticulum stress and sensitizes sarcoma cells toward

192. Kojima K, Konopleva M, Samudio IJ, et al. Concomitant inhibition of MDM2 and


Bcl-2 protein function synergistically induce mitochondrial apoptosis in AML. Cell Cycle 2006;5(23):2778-86 193. Korcsmáros T, Szalay M, Böde C, et al. How to design multi-target drugs: search options in cellular networks. Expert Opin Drug Discov 2007;2(6):799-808 194. Elliott PJ, Soucy TA, Pien CS, et al. Assays for proteasome inhibition. Methods Mol Med 2003;85:163-72 195. Lagoja I, Herdewijn P. Use of RNA in drug design. Expert Opin Drug Discov 2007;2(6):889-903 196. Woo KJ, Lee TJ, Lee SH, et al. Elevated gadd153/chop expression during resveratrol-induced apoptosis in human colon cancer cells. Biochem Pharmacol 2007;73(1):68-76 197. Sanges D, Marigo V. Cross-talk between two apoptotic pathways activated by endoplasmic reticulum stress: Differential contribution of caspase-12 and AIF. Apoptosis 2006;11(9):1629-41

Afﬁliation Donavon C Hiss†1 MSc (Med) PhD (Med) & Gary A Gabriels2 MSc (Med) †Author for correspondence 1Head, Molecular Oncology Research Programme, University of the Western Cape, Department of Medical BioSciences, Bellville, 7535, South Africa Tel: +27 21 959 2334; Fax: +27 959 1563; E-mail: [email protected] 2Chief Medical Scientific Officer, University of Cape Town, Observatory, Division of Clinical Pharmacology, Department of Medicine, 7925, South Africa

921