Current Cancer Drug Targets - Volume 11, Issue 3, 2011

Dysregulated cellular proliferation and apoptotic pathways are hallmarks of human cancers. In recent years, the ubiquitinproteasome system has been identified as essential for maintaining the cell growth-death balance and for the development and progression of several human malignancies. Emerging evidence demonstrates that targeting the tumor ubiquitin-proteasome degradation pathway is a promising strategy for cancer treatment. The approval of bortezomib by the US Food and Drug Administration in 2003 represented a significant milestone as the first proteasome inhibitor used to treat cancer. Other novel proteasome inhibitors, such as marizomib and carfilzomib, have also been developed and are being tested in clinical trials. Also, some natural products with proteasome-inhibitory effects, such as green tea polyphenols and other dietary flavonoids, have been studied alone and in combination with traditional chemotherapy and radiotherapy against various cancers. Furthermore, some old copper-binding drugs such as clioquinol and disulfiram were shown to inhibit the proteasome activity in human cancer cells in vitro and in vivo, demonstrating new uses for old drugs. E3 ligases have also recently been identified as potential cancer targets and biomarkers, because ubiquitin-proteasome pathway is mainly carried out by substrate-specific E3 ligases, which are dysregulated in human cancers. This special issue focuses on compounds, new and old, that can target the ubiquitin-proteasome pathway in human cancers. This issue also presents summaries of the latest outstanding developments in the medicinal chemistry, pharmacology, molecular biology and biochemistry of these selected anticancer drugs and agents. The authors of the first article [1] summarize current knowledge on bortezomib, the first proteasome inhibitor anticancer drug used clinically. Based on positive preclinical and clinical studies, bortezomib was approved for clinical use as a front-line treatment for newly diagnosed multiple myeloma patients and for the treatment of relapsed/refractory multiple myeloma and mantle cell lymphoma, for which this drug has become the staple of treatment. Both by itself and in combination therapies, bortezomib has benefited patients in these clinical studies by inducing tumor cell death or chemo-/radio-sensitization and overcoming drug resistance in hematological malignancies. However, its effect on solid tumors has been less than encouraging. Additionally, the widespread clinical use of bortezomib has been hampered by the appearance of dose-limiting toxicities, drugresistance and interference with some natural compounds. These findings have encouraged scientists to develop next generation proteasome inhibitors that broaden the spectrum of efficacy and produce a more durable clinical response in cancer patients. In this vein, the second article summarizes the preclinical profile and clinical trials of marizomib, a member of a new generation of proteasome inhibitor anticancer drugs [2]. The potent and sustained inhibition of all three proteolytic activities of the proteasome by marizomib has inspired extensive preclinical evaluation in a variety of hematologic and solid tumor models, where it has shown efficacy as a single agent and in combination with biologics, chemotherapeutics and targeted therapeutic agents. These studies provided the framework for ongoing clinical trials in patients with multiple myeloma, lymphomas, leukemias and solid tumors, including those who have failed bortezomib treatment, as well as in patients where other proteasome inhibitors have failed to demonstrate significant efficacy [2]. Two other second generation proteasome inhibitors, carfilzomib and immunoproteasome-specific inhibitors (IPSIs) are reviewed in the third article [3]. As an irreversible proteasome inhibitor, carfilzomib has shown preclinical effectiveness against hematological and solid malignancies both in vitro and in vivo. Carfilzomib is also currently achieving successful response rates within the clinical setting. In addition to conventional proteasome inhibitors, a novel approach may be to specifically target the hematological-specific immunoproteasome, thereby increasing overall effectiveness and reducing deleterious off-target effects. The immunoproteasome-specific inhibitor IPSI-001 was shown to exhibit inhibitory preference over the constitutive proteasome, and display enhanced efficacy toward apoptotic induction of tumor cells from a hematologic origin [3]. While the discovery of proteasome inhibitors as anticancer drugs has validated the concept of the proteasome as a novel cancer drug target, researchers have also been looking for natural compounds that could selectively inhibit the proteasome activity in human tumor cells. Epigallocatechin-3-gallate (EGCG) is the most abundant and active compound in green tea. It is extensively studied for its cancer-preventive and anticancer activities as well as its cellular targets. One molecular target of EGCG is the proteasome [4]. The fourth review article discusses the potential use of green tea polyphenols as proteasome inhibitors for chemoprevention and cancer therapies. The authors also describe a synthetic prodrug of EGCG that has improved bioavailability, stability, and proteasome-inhibitory activities against various human cancer cells and tumors compared to natural EGCG [4]. The proteasome-inhibiting ability of other dietary flavonoids and naturally occurring compounds is the topic of the fifth review article [5]. The authors discuss recent advances in proteasomal modulation by some naturally occurring polyphenols, including curcumin, quercetin, apigenin, and resveratrol, and by several natural product extracts. They provide evidence that the proteasome is a viable therapeutic target for cancer prevention and treatment. The authors also discuss the potential interaction of bortezomib with some natural compounds which can reverse the anticancer effects of bortezomib and prevent malignant cells from undergoing apoptosis. Recent studies have integrated the fields of proteasome inhibitors and anti-copper drugs. The sixth review article discusses the potential of clioquinol, an old copper-binding drug, as a novel copper-dependent and -independent proteasome inhibitor [6]. Clioquinol displays preclinical efficacy in the treatment of malignancy. Its anticancer activity is due, at least in part, to its ability to inhibit the proteasome through mechanisms dependent and independent of its ability to bind heavy metals such as copper. Thus, clioquinol represents a novel therapeutic strategy to inhibit the proteasome. Given prior toxicology and pharmacological studies, clioquinol could be rapidly repositioned for a new anticancer indication [6]. Targeting the proteasome pathway in malignancies with disulfiram, a clinically used anti-alcoholism drug that binds copper, is discussed in the next review article [7]. Disulfiram has been used for decades in alcohol aversion therapy, and its metabolite dithiocarb was shown in the 1970s to suppress cancer growth in vivo and even in human patients. This drug is able to target the proteasome, which is involved in multidrug resistance, tumor angiogenesis and invasion. The author also discusses ongoing clinical trials with disulfiram as an adjuvant therapy against lung cancer and as a monotherapy against liver metastasis [7]. The next article further reviews the status of disulfiram and disulfiram derivatives as novel potential anticancer drugs targeting the ubiquitin-proteasome system in both preclinical and clinical studies [8]. Despite data from the 1970s and 80s showing that disulfiram and analogs are able to enhance the activity of anticancer cytotoxic drugs and may be useful as chemopreventative agents, the underlying molecular mechanisms have remained elusive until recently. Large scale screening efforts revealed that disulfiram has proteasome-inhibitory activity. Moreover, disulfiram was also found to have specific activity against zinc fingers and RING-finger ubiquitin E3 ligases that play an important role in cancer development [8]. The SCF (Skp1-Cullin-F-box protein) multisubunit complex is the largest E3 ubiquitin ligase family that promotes the ubiquitination of various regulatory proteins for targeted degradation, thus regulating many biological processes, including cell cycle progression, signal transduction, and DNA replication. The last review article in this issue describes recent advances in validation of SCF E3 ubiquitin ligase complexes as an attractive anticancer target and discusses how SCF E3 ligase inhibitors have been developed into a novel class of anticancer agents [9]....

Targeting the ubiquitin-proteasome pathway has emerged as a rational approach in the treatment of human cancer. Based on positive preclinical and clinical studies, bortezomib was subsequently approved for the clinical use as a front-line treatment for newly diagnosed multiple myeloma patients and for the treatment of relapsed/refractory multiple myeloma and mantle cell lymphoma, for which this drug has become the staple of treatment. The approval of bortezomib by the US Food and Drug Administration (FDA) represented a significant milestone as the first proteasome inhibitor to be implemented in the treatment of malignant disease. Bortezomib has shown a positive clinical benefit either alone or as a part of combination therapy to induce chemo-/radio-sensitization or overcome drug resistance. One of the major mechanisms of bortezomib associated with its anticancer activity is through upregulation of NOXA, which is a proapoptotic protein, and NOXA may interact with the anti-apoptotic proteins of Bcl-2 subfamily Bcl-XL and Bcl-2, and result in apoptotic cell death in malignant cells. Another important mechanism of bortezomib is through suppression of the NF-κB signaling pathway resulting in the down-regulation of its anti-apoptotic target genes. Although the majority of success achieved with bortezomib has been in hematological malignancies, its effect toward solid tumors has been less than encouraging. Additionally, the widespread clinical use of bortezomib continues to be hampered by the appearance of dose-limiting toxicities, drug-resistance and interference by some natural compounds. These findings could help guide physicians in refining the clinical use of bortezomib, and encourage basic scientists to generate next generation proteasome inhibitors that broaden the spectrum of efficacy and produce a more durable clinical response in cancer patients. Other desirable applications for the use of proteasome inhibitors include the development of inhibitors against specific E3 ligases, which act at an early step in the ubiquitin-proteasome pathway, and the discovery of less toxic and novel proteasome inhibitors from natural products and traditional medicines, which may provide more viable drug candidates for cancer chemoprevention and the treatment of cancer patients in the future.

The proteasome has emerged as an important clinically relevant target for the treatment of hematologic malignancies. Since the Food and Drug Administration approved the first-in-class proteasome inhibitor bortezomib (Velcade® ) for the treatment of relapsed/refractory multiple myeloma (MM) and mantle cell lymphoma, it has become clear that new inhibitors are needed that have a better therapeutic ratio, can overcome inherent and acquired bortezomib resistance and exhibit broader anti-cancer activities. Marizomib (NPI-0052; salinosporamide A) is a structurally and pharmacologically unique β-lactone-γ-lactam proteasome inhibitor that may fulfill these unmet needs. The potent and sustained inhibition of all three proteolytic activities of the proteasome by marizomib has inspired extensive preclinical evaluation in a variety of hematologic and solid tumor models, where it is efficacious as a single agent and in combination with biologics, chemotherapeutics and targeted therapeutic agents. Specifically, marizomib has been evaluated in models for multiple myeloma, mantle cell lymphoma, Waldenstrom's macroglobulinemia, chronic and acute lymphocytic leukemia, as well as glioma, colorectal and pancreatic cancer models, and has exhibited synergistic activities in tumor models in combination with bortezomib, the immunomodulatory agent lenalidomide (Revlimid® ), and various histone deacetylase inhibitors. These and other studies provided the framework for ongoing clinical trials in patients with MM, lymphomas, leukemias and solid tumors, including those who have failed bortezomib treatment, as well as in patients with diagnoses where other proteasome inhibitors have not demonstrated significant efficacy. This review captures the remarkable translational studies and contributions from many collaborators that have advanced marizomib from seabed to bench to bedside.

The ubiquitin-proteasome pathway (UPP) is an attractive chemotherapeutic target due to its intrinsically stringent regulation of cell cycle, pro-survival, and anti-apoptotic regulators that disproportionately favor survival and proliferation in malignant cells. A reversible first-in-class proteasome inhibitor, bortezomib, is Food and Drug Administration approved for multiple myeloma and relapsed/refractory mantle cell lymphoma and has proven to be extremely effective, both as a single agent and in combination. An irreversible second generation proteasome inhibitor, carfilzomib, has shown preclinical effectiveness against hematological and solid malignancies both in vitro and in vivo. Carfilzomib, a peptidylepoxyketone functions similarly to bortezomib through primary inhibition of chymotrypsin-like (ChT-L) activity at the β5 subunits of the core 20S proteasome. Carfilzomib is also currently achieving successful response rates within the clinical setting. In addition to conventional proteasome inhibitors, a novel approach may be to specifically target the hematological- specific immunoproteasome, thereby increasing overall effectiveness and reducing negative off-target effects. The immunoproteasome-specific inhibitor, IPSI-001, was shown to have inhibitory preference over the constitutive proteasome, and display enhanced efficiency of apoptotic induction of tumor cells from a hematologic origin. Herein, we discuss the preclinical and clinical development of carfilzomib and explore the potential of immunoproteasome-specific inhibitors, like IPSI-001, as a rational approach to exclusively target hematological malignancies.

Next to water, tea is the most popular beverage in the world. The most abundant and active compound in green tea is (-)-epigallocatechin-3-gallate (EGCG), which is extensively studied for its cancer-preventive and anti-cancer activities as well as its cellular targets. One potential molecular target of EGCG is the proteasome. While molecular docking and structure-activity relationship (SAR) analysis suggests that the ester carbon of EGCG is important for mediating its proteasome-inhibitory activity, EGCG is very unstable under physiological conditions. Therefore, a series of analogs were synthesized aiming to improve stability and bioavailability of EGCG. Among them, peracetate-protected or the prodrug of EGCG was found to have increased bioavailability, stability, and proteasome-inhibitory activities against various human cancer cells and tumors compared to EGCG, suggesting its potential use for cancer prevention and treatment. Epidemiological studies have indicated that green tea consumption is associated with the reduced risk of cancers, especially associated with the reduced risk of late stage of cancers. This risk reduction may be attributed not only to proteasome inhibition, but also to numerous other intracellular molecules targeted by EGCG that are involved in cell cycle regulation, apoptosis, angiogenesis, and metastasis.

Aberrant cellular proliferation and compromised apoptotic pathways are hallmarks of cancer aggressiveness, and in this framework, the role of protein degradation machineries have been extensively dissected. Among proteases, the proteasome is unequivocally central in the intracellular regulation of both these processes, thus several proteasome-directed therapies have been investigated, aiming at controlling its activity and possibly restoring normal cell functions. Numerous studies reported proteasome inhibitors (both synthetic and natural occurring) to potently and selectively induce apoptosis in many types of cancer cells. In this review, we discuss recent advances in proteasomal modulation by some natural occurring polyphenols, globally providing evidence of the proteasome role as therapeutic target in cancer treatment.

Clioquinol (5-chloro-7-iodo-quinolin-8-ol) was used in the 1950's-1970's as an oral anti-parasitic agent. More recently, studies have demonstrated that Clioquinol displays preclinical efficacy in the treatment of malignancy. Its anticancer activity relates, at least in part, to its ability to inhibit the proteasome through mechanisms dependent and independent of its ability to bind heavy metals such as copper. By acting as a metal ionophore Clioquinol transports metal ions from the extracellular environment into the cell and mobilizes weakly bound intracellular stores. It then directs the metal to the proteasome resulting in disruption of this enzymatic complex. In addition, Clioquinol is capable of directly inhibiting the proteasome at higher concentrations. Thus, Clioquinol represents a novel therapeutic strategy to inhibit the proteasome. Given the prior toxicology and pharmacology studies, Clioquinol could be rapidly repositioned for a new anticancer indication. This review highlights the mechanism of action of Clioquinol as a proteasome inhibitor. In addition, it discusses the human pharmacology and toxicology studies and how this information would guide a phase I clinical trial of this agent for patients with malignancy.

An old drug, Antabuse (disulfiram), used for decades in alcohol aversion therapy, and its metabolite Ditiocarb were shown from 1970s to suppress cancer growth in vivo and even in human patients. The drug targets multidrug resistance, angiogenesis, invasion, and proteasome. Today, there are ongoing clinical trials of Antabuse as an adjuvant therapy against lung cancer and as a monotherapy against cancers metastasizing to liver. The larger clinical trials, if appropriate, will need support from governments and charities to get the generic drug into the clinic as a “non-profit” drug.

Disulfiram is a FDA approved drug for the treatment of alcoholism and has been available for clinical use for over five decades. Despite data from the 1970s and 80s, which showed that disulfiram and analogs are able to enhance the activity of anticancer cytotoxic drugs and might be useful as chemopreventative agents, the underlying molecular mechanisms remained unknown until recently. Large scale screening efforts for agents that can inhibit the proteasome and be used as novel anticancer drugs revealed that disulfiram has proteasome inhibitory activity. Moreover, disulfiram was also found to have specific activity against zinc fingers and RING-finger ubiquitin E3 ligases that play an important role in cancer development. Here we review the preclinical and clinical studies exploring disulfiram as an anticancer agent, as well as research programs that focus on the development of disulfiram derivatives as inhibitors of the ubiquitinproteasome system.

The SCF (Skp1, Cullins, F-box proteins) multisubunit E3 ubiquitin ligase, also known as CRL (Cullin-RING ubiquitin Ligase) is the largest E3 ubiquitin ligase family that promotes the ubiquitination of various regulatory proteins for targeted degradation, thus regulating many biological processes, including cell cycle progression, signal transduction, and DNA replication. The efforts to discover small molecule inhibitors of a SCF-type ligase or its components were expedited by the FDA approval of Bortezomib (also known as Velcade or PS-341), the first (and only) class of general proteasome inhibitor, for the treatment of relapsed/refractory multiple myeloma and mantle cell lymphoma. Although Bortezomib has demonstrated a certain degree of cancer cell selectivity with measurable therapeutic index, the drug is, in general, cytotoxic due to its inhibition of overall protein degradation. An alternative and ideal approach is to target a specific E3 ligase, known to be activated in human cancer, for a high level of specificity and selectivity with less associated toxicity, since such inhibitors would selectively stabilize a specific set of cellular proteins regulated by this E3. Here, we review recent advances in validation of SCF E3 ubiquitin ligase as an attractive anti-cancer target and discuss how MLN4924, a small molecule inhibitor of NEDD8-activating enzyme, can be developed as a novel class of anticancer agents by inhibiting SCF E3 ligase via removal of cullin neddylation. Finally, we discuss under future perspective how basic research on SCF biology will direct the drug discovery efforts surrounding this target.

Autophagy is a catabolic process whereby cells maintain homeostasis by eliminating unnecessary proteins and damaged organelles. It may be triggered under physiological conditions, such as nutrient starvation, or in response to a variety of stress stimuli, such as exposure to radiations or cytotoxic compounds. Although autophagy is basically a protective mechanism that sustains cell survival under adverse conditions, it has been recently demonstrated that the induction of autophagic process may ultimately lead to cell death. As for the role of autophagy in cancer, it is still very controversial whether it suppresses tumorigenesis or provides cancer cells with a rescue mechanism under unfavourable conditions. Therefore, the dual role of autophagy in tumor progression and in the response of cancer cells to chemotherapeutic drugs is still open to debate. The first part of this review describes the cellular events occurring during the various phases of the autophagic process. Special attention has been given to the morphological aspects and the regulatory molecules involved in autophagic cell death. Specifically, we have focused on the proteins necessary for autophagosome formation, encoded by the ATG (AuTophaGy-related gene) gene family, and their role in the regulation of the process of autophagy. We also examined the effects of autophagy modulators on cell survival and cell death and discussed the recent efforts aimed at finding novel agents that activate or inhibit autophagy by targeting regulatory molecules of the complex autophagy pathways.

The peptidyl prolyl isomerase (Pin1) that induces cis-trans isomerization of the peptide bond involving serine/ threonine-proline has recently been shown to regulate the activity of many phosphoproteins including the ones involved in damage response pathways. We investigated Pin1 as a potential target for enhancing the efficacy of anticancer therapy by studying the effects of juglone, a Pin1 inhibitor on the cytotoxicity of etoposide (a widely used anticancer drug that targets topoisomerase IIα) in human tumor cell lines. Treatment of cells with juglone synergistically enhanced the cytotoxicity of etoposide (loss of clonogenicity) with a tenfold increase when etoposide treatment preceded juglone exposure. On the other hand, the toxicity was less than additive when the treatment protocol was reversed (i.e exposure to juglone followed by etoposide treatment). This suggests that Pin1 inhibition possibly reduces the induction of initial DNA damage by etoposide, which was supported by a decrease in the levels of chromatin bound topoIIα. Increase in the etoposide induced toxicity by juglone appeared to be mainly due to enhanced mitotic cell death linked to cytogenetic damage, although a moderate increase in interphase (apoptotic) death was also evident as revealed by DNA degradation (hypodiploid population and TUNEL assay). Since the level of Pin1 is found to be higher in cancer cells, this enzyme could be a potential target for developing an adjuvant to enhance the efficacy of anticancer therapies.