STAT5A/B Blockade Sensitizes Prostate Cancer to Radiation through Inhibition of RAD51 and DNA Repair.

Purpose: The standard treatment for organ-confined prostate cancer is surgery or radiation, and locally advanced prostate cancer is typically treated with radiotherapy alone or in combination with androgen deprivation therapy. Here, we investigated whether Stat5a/b participates in regulation of double-strand DNA break repair in prostate cancer, and whether Stat5 inhibition may provide a novel strategy to sensitize prostate cancer to radiotherapy.Experimental Design: Stat5a/b regulation of DNA repair in prostate cancer was evaluated by comet and clonogenic survival assays, followed by assays specific to homologous recombination (HR) DNA repair and nonhomologous end joining (NHEJ) DNA repair. For HR DNA repair, Stat5a/b regulation of Rad51 and the mechanisms underlying the regulation were investigated in prostate cancer cells, xenograft tumors, and patient-derived prostate cancers ex vivo in 3D explant cultures. Stat5a/b induction of Rad51 and HR DNA repair and responsiveness to radiation were evaluated in vivo in mice bearing prostate cancer xenograft tumors.Results: Stat5a/b is critical for Rad51 expression in prostate cancer via Jak2-dependent mechanisms by inducing Rad51 mRNA levels. Consistent with this, genetic knockdown of Stat5a/b suppressed HR DNA repair while not affecting NHEJ DNA repair. Pharmacologic Stat5a/b inhibition potently sensitized prostate cancer cell lines and prostate cancer tumors to radiation, while not inducing radiation sensitivity in the neighboring tissues.Conclusions: This work introduces a novel concept of a pivotal role of Jak2-Stat5a/b signaling for Rad51 expression and HR DNA repair in prostate cancer. Inhibition of Jak2-Stat5a/b signaling sensitizes prostate cancer to radiation and, therefore, may provide an adjuvant therapy for radiation to reduce radiation-induced damage to the neighboring tissues. Clin Cancer Res; 24(8); 1917-31. ©2018 AACR.

Involvement of ALAD-20S Proteasome Complexes in Ubiquitination and Acetylation of Proteasomal α2 Subunits.

The ubiquitin-proteasome pathway has gained attention as a potential chemotherapeutic target, owing to its importance in the maintenance of protein homeostasis and the observation that cancer cells are more dependent on this pathway than normal cells. Additionally, inhibition of histone deacetylases (HDACs) by their inhibitors like Vorinostat (SAHA) has also proven a useful strategy in cancer therapy and the concomitant use of proteasome and HDAC inhibitors has been shown to be superior to either treatment alone. It has also been reported that delta-aminolevulinic acid dehydratase (ALAD) is a proteasome-associated protein, and may function as an endogenous proteasome inhibitor. While the role of ALAD in the heme biosynthetic pathway is well characterized, little is known about its interaction with, and the mechanism by which it inhibits, the proteasome. In the present study, this ALAD-proteasome complex was further characterized in cultured prostate cancer cells and the effects of SAHA treatment on the regulation of ALAD were investigated. ALAD interacts with the 20S proteasomal core, but not the 19S regulatory cap. Some ubiquitinated species were detected in ALAD immunoprecipitates that have similar molecular weights to ubiquitinated proteasomal α2 subunits, suggesting preferred binding of ALAD to ubiquitinated α2. Additionally, SAHA treatment increases levels of ALAD protein and an acetylated protein with a molecular weight similar to the ubiquitinated α2 subunit. Thus, the results of this study suggest that ALAD may play a regulatory role in a previously unreported post-translational modification of proteasomal α subunits.

Metal-based 2,3-indolinedione derivatives as proteasome inhibitors and inducers of apoptosis in human cancer cells.

Proliferation and apoptotic pathways are tightly regulated in cells by the ubiquitin-proteasome system (UPS). Alterations in the UPS may result in cellular transformation or other pathological conditions. The proteasome is indeed often found to be overactive in cancer cells. It has been reported that 2,3-indolinedione (L), which exists in marine organisms, as well as in mammals, is a proteasome inhibitor. Studies have shown that metal-based complexes inhibit proteasome activity and induce apoptosis in certain human cancer cells. In the current study, we synthesized six novel metal-based complexes with derivatives of 2,3-indolinedione: [Cd (C15H11O3N2) (CH3COO)] (C1), [Cd (C15H11O2N2) (CH3COO)] (C2), [Co (C15H9O4N2) (CH3COO)] (C3), [Co (C15H11O2N2) (CH3COO)] (C4), [Zn (C19H14O3N3) (CH3COO)] (C5) and [Zn (C17H13O3N2) (CH3COO)] (C6). We sought to characterize and assess the proteasome inhibitory and anti-proliferative effects of these metal-based complexes in human breast (MDA-MB-231) and prostate (LNCaP and PC-3) cancer cells, in order to determine whether specific structures contribute to the inhibition of tumor proteasome activity and the induction of apoptosis. The results revealed that the complexes, C1, C3 and C5, but not their counterparts, C2, C4 and C6, inhibited the chymotrypsin-like activity of the human cancer cellular 26S proteasome; in addition, these complexes promoted the accumulation of the proteasome target protein, Bax, inhibited cell growth and induced apoptosis in a concentration- and time-dependent manner due to their unique structures. Our data suggest that the study of metal-based complexes, including aromatic ring structures with electron-attracting groups, may be an interesting research direction for the development of anticancer drugs.

Gold(III)-dithiocarbamato peptidomimetics in the forefront of the targeted anticancer therapy: preclinical studies against human breast neoplasia.

Since the serendipitous discovery of cisplatin, platinum-based drugs have become well-established antitumor agents, despite the fact that their clinical use is limited by many severe side-effects. In order to both improve the chemotherapeutic index and broaden the therapeutic spectrum of current drugs, our most recent anti-neoplastic agents, Au(III) complexes, were designed as carrier-mediated delivery systems exploiting peptide transporters, which are up-regulated in some cancers. Among all, we focused on two compounds and tested them on human MDA-MB-231 (resistant to cisplatin) breast cancer cell cultures and xenografts, discovering the proteasome as a major target both in vitro and in vivo. 53% inhibition of breast tumor growth in mice was observed after 27 days of treatment at 1.0 mg kg(-1) d(-1), compared to control. Remarkably, if only the most responsive mice are taken into account, 85% growth inhibition, with some animals showing tumor shrinkage, was observed after 13 days. These results led us to file an international patent, recognizing this class of gold(III) peptidomimetics as suitable candidates for entering phase I clinical trials.

Chemotherapeutic inhibitors in the treatment of prostate cancer.

INTRODUCTION: Prostate cancer being the second leading cause of death in men in Western countries remains a major challenge in healthcare. Several novel agents targeting signaling pathways in prostate cancer have recently been approved by the US Food and Drug Administration (FDA) but there is still an unmet need for new treatment strategies for castration-resistant prostate cancer (CRPC). AREAS COVERED: This review provides a broad overview of prostate cancer therapeutics and highlights key players in the biology of prostate cancer as well as first- and second-line treatments for CRPC. Keywords 'chemotherapeutic agents', 'prostate cancer', 'Phase III clinical trials' and 'US FDA approval' were used for search in PubMed and clinicalTrials.gov databases and the obtained literature was reviewed and summarized. EXPERT OPINION: Owing to the advances in screening and diagnostic techniques, the majority of prostate cancer cases are diagnosed at an early stage resulting in an almost 100% 5-year survival rate. Recently FDA-approved novel agents (e.g., abiraterone acetate and enzalutamide) have provided new hope in the fight against prostate cancer. However, CRPC remains an incurable disease. Identification of mechanisms of resistance, new biomarkers, appropriate clinical trial end points and novel treatments holds the key for the future of prostate cancer therapy.

1,10-Phenanthroline promotes copper complexes into tumor cells and induces apoptosis by inhibiting the proteasome activity.

Indole-3-acetic acid and indole-3-propionic acid, two potent natural plant growth hormones, have attracted attention as promising prodrugs in cancer therapy. Copper is known to be a cofactor essential for tumor angiogenesis. We have previously reported that taurine, L-glutamine, and quinoline-2-carboxaldehyde Schiff base copper complexes inhibit cell proliferation and proteasome activity in human cancer cells. In the current study, we synthesized two types of copper complexes, dinuclear complexes and ternary complexes, to investigate whether a certain structure could easily carry copper into cancer cells and consequently inhibit tumor proteasome activity and induce apoptosis. We observed that ternary complexes binding with 1,10-phenanthroline are more potent proteasome inhibitors and apoptosis inducers than dinuclear complexes in PC-3 human prostate cancer cells. Furthermore, the ternary complexes potently inhibit proteasome activity before induction of apoptosis in MDA-MB-231 human breast cancer cells, but not in nontumorigenic MCF-10A cells. Our results suggest that copper complexes binding with 1,10-phenanthroline as the third ligand could serve as potent, selective proteasome inhibitors and apoptosis inducers in tumor cells, and that the ternary complexes may be good potential anticancer drugs.

Novel Polypyridyl chelators deplete cellular zinc and destabilize the X-linked inhibitor of apoptosis protein (XIAP) prior to induction of apoptosis in human prostate and breast cancer cells.

X-linked inhibitor of apoptosis protein (XIAP), inhibits the initiation and execution phases of the apoptotic pathway. XIAP is the most potent member of the inhibitor of apoptosis protein (IAP) family of the endogenous caspase inhibitors. Therefore, targeting XIAP may be a promising strategy for the treatment of apoptosis-resistant malignancies. In this study, we systematically studied the relationships of chemical structures of several novel ligands to their zinc (Zn)-binding ability, molecular target XIAP, and tumor cell death-inducing activity. We show that treatment of PC-3 prostate cancer and MDA-MB-231 breast cancer cells with these membrane-permeable Zn-chelators with different Zn affinities results in varying degrees of XIAP depletion. Following decreased level of XIAP expression, we also show apoptosis-related caspase activation and cellular morphological changes upon treatment with strong Zn-chelators N4Py and BnTPEN. Addition of Zn has a full protective effect on the cells treated with these chelators, while iron (Fe) addition has only partial protection that, however, can be further increased to a comparable level of protection as Zn by inhibition of ROS generation, indicating that cell death effects mediated by Fe- but not Zn-complexes involve redox cycling. These findings suggest that strong Zn-chelating agents may be useful in the treatment of apoptosis-resistant human cancers.

New applications of old metal-binding drugs in the treatment of human cancer.

Significant advances in the use of metal complexes, precipitated by platinum, have fostered a renewed interest in harnessing their rich potential in the treatment of cancer. In addition to platinum-based complexes, the anticancer properties of other metals such as ruthenium have been realized, and ruthenium-based compounds are currently being investigated in clinical trials. Since the process of drug development can be expensive and cumbersome, finding new applications of existing drugs may provide effective means to expedite the regulatory process in bringing new drugs to the clinical setting. Encouraging findings from laboratory studies reveal significant anticancer activity from different classes of metal-chelating compounds, such as disulfiram, clioquinol, and dithiocarbamate derivatives that are currently approved for the treatment of various pathological disorders. Their use as coordination complexes with metals such as copper, zinc, and gold that target the ubiquitin-proteasome pathway have shown significant promise as potential anticancer agents. This review discusses the unique role of several selected metals in relation to their anti-cancer properties as well as the new therapeutic potential of several previously approved metal-chelating drugs. In vitro and in vivo experimental evidence along with mechanisms of action (e.g., via targeting the tumor proteasome) will also be discussed with anticipation of strengthening this exciting new concept.

Inhibition of the purified 20S proteasome by non-heme iron complexes.

Polypyridyl pentadentate ligands N4Py (1) and Bn-TPEN (2), along with their respective iron complexes, have been investigated for their ability to inhibit the purified 20S proteasome. Results demonstrated that the iron complexes of both ligands are potent inhibitors of the 20S proteasome (IC(50) = 9.2 µM for [Fe(II)(OH(2))(N4Py)](2+) (3) and 4.0 µM for [Fe(II)(OH(2))(Bn-TPEN)](2+) (4)). Control experiments showed that ligand 1 or Fe(II) alone showed no inhibition, whereas 2 was moderately active (IC(50) = 96 µM), suggesting that iron, when bound to these ligands, plays a key role in proteasome inhibition. Results from time-dependent inactivation studies suggest different modes of action for the iron complexes. Time-dependent decay of proteasome activity was observed upon incubation in the presence of 4, which accelerated in the presence of DTT, suggesting reductive activation of O(2) and oxidation of the 20S proteasome as a mode of action. In contrast, loss of 20S proteasome activity was not observed with 3 over time, suggesting inhibition through direct binding of the iron complex to the enzyme. Inhibition of the 20S proteasome by 4 was not blocked by reactive oxygen species scavengers, consistent with a unique oxidant being responsible for the time-dependent inhibition observed.