Lawrence N. Barrera

BTK inhibitor ibrutinib is cytotoxic to myeloma and potently enhances bortezomib and lenalidomide activities through NF-κB

Ibrutinib (previously known as PCI-32765) has recently shown encouraging clinical activity in chronic lymphocytic leukaemia (CLL) effecting cell death through inhibition of Brutons tyrosine kinase (BTK). In this study we report for the first time that ibrutinib is cytotoxic to malignant plasma cells from patients with multiple myeloma (MM) and furthermore that treatment with ibrutinib significantly augments the cytotoxic activity of bortezomib and lenalidomide chemotherapies. We describe that the cytotoxicity of ibrutinib in MM is mediated via an inhibitory effect on the nuclear factor-κB (NF-κB) pathway. Specifically, ibrutinib blocks the phosphorylation of serine-536 of the p65 subunit of NF-κB, preventing its nuclear translocation, resulting in down-regulation of anti-apoptotic proteins Bcl-xL, FLIP(L) and survivin and culminating in caspase-mediated apoptosis within the malignant plasma cells. Taken together these data provide a platform for clinical trials of ibrutinib in myeloma and a rationale for its use in combination therapy, particularly with bortezomib.

Epigenetic and antioxidant effects of dietary isothiocyanates and selenium: potential implications for cancer chemoprevention.

There is evidence from epidemiological studies suggesting that increased consumption of cruciferous vegetables may protect against specific cancers more effectively than total fruit and vegetable intake. These beneficial effects are attributed to the glucosinolate breakdown products, isothiocyanates (ITC). Similarly, selenium (Se) consumption has also been inversely associated with cancer risk and as an integral part of many selenoproteins may influence multiple pathways in the development of cancer. This paper will briefly review the current state of knowledge concerning the effect of Se and ITC in cancer development with a particular emphasis on its antioxidant properties, and will also address whether alterations in DNA methylation may be a potential mechanism whereby these dietary constituents protect against the carcinogenic process. Furthermore, we will discuss the advantages of combining ITC and Se to benefit from their complementary mechanisms of action to potentially protect against the alterations leading to neoplasia. Based on this review it may be concluded that an understanding of the impact of ITC and Se on aberrant DNA methylation in relation to factors modulating gene-specific and global methylation patterns, in addition to the effect of these food constituents as modulators of key selenoenzymes, such as gastrointestinal glutathione peroxidase-2 (GPx2) and thioredoxin reductase-1 (TrxR1), may provide insights into the potential synergy among various components of a plant-based diet that may counteract the genetic and epigenetic alterations that initiate and sustain neoplasia.

Bortezomib induces heme oxygenase-1 expression in multiple myeloma

Multiple myeloma (MM) is a progressive malignant disorder characterized by accumulation of plasma cells in the bone marrow. MM remains an incurable disease with a 5-y survival rate of approximately 40%. While clinical response rates to first line chemotherapeutics are high, disease relapse is inevitable, and occurs because a small fraction of the original myeloma cells appear to be resistant to treatment. Heme oxygenase-1 (HO-1) is an Nrf2 transcription factor-regulated gene that is commonly induced following oxidative stress and cellular injury, functioning to decrease oxidative stress and inflammatory responses, protecting against apoptosis and altering the cell cycle. We and others have highlighted the role of HO-1 in providing cellular protection against chemotherapeutic drugs in a number of cancer cells, which we have highlighted here in this Extra View. Furthermore, we explored the expression of HO-1 in multiple myeloma cells in response to the key anti-myeloma drugs bortezomib and lenalidomide. We show here for the first time that bortezomib increases HO-1 expression in a time- and concentration-dependent manner. Moreover, we also observe that HO-1 is increased in lenalidomide-resistant MM cell lines. Altogether, we highlight a possible role for HO-1 in basal and acquired chemoresistance in MM.

Synergy between sulforaphane and selenium in the up-regulation of thioredoxin reductase and protection against hydrogen peroxide-induced cell death in human hepatocytes

Dietary isothiocyanates and selenium are chemopreventive agents and potent inducers of antioxidant enzymes. It has been previously shown that sulforaphane and selenium have a synergistic effect on the upregulation of thioredoxin reductase-1 (TrxR-1) in human hepatoma HepG2 cells. In this paper, further evidence is presented to show that sulforaphane and selenium synergistically induce TrxR-1 expression in immortalised human hepatocytes. Sulforaphane was found to be more toxic toward hepatocytes than HepG2 cells with IC50=25.1 and 56.4 μM, respectively. Sulforaphane can protect against hydrogen peroxide-induced cell death and this protection was enhanced by co-treatment with selenium. Using siRNA to knock down TrxR-1 or Nrf2, sulforaphane (5 μM)-protected cell viability was reduced from 73% to 46% and 34%, respectively, suggesting that TrxR-1 is an important enzyme in protection against hydrogen peroxide-induced cell death. Sulforaphane-induced TrxR-1 expression was positively associated with significant levels of Nrf2 translocation into the nucleus, but co-treatment with selenium showed no significant increase in Nrf2 translocation. Moreover, MAPK (ERK, JNK and p38) and PI3K/Akt signalling pathways were found to play no significant role in sulforaphane-induced Nrf2 translocation into the nucleus. However, blocking ERK and JNK signalling pathways decreased sulforaphane-induced TrxR-1 mRNA by about 20%; whereas blocking p38 and PI3K/AKT increased TrxR-1 transcription. In summary, a combination of sulforaphane and selenium resulted in a synergistic upregulation of TrxR-1 that contributed to the enhanced protection against free radical-mediated oxidative damage in human hepatocytes.

TrxR1 and GPx2 are potently induced by isothiocyanates and selenium, and mutually cooperate to protect Caco-2 cells against free radical-mediated cell death.

Currently, there is significant interest in the field of diet-gene interactions and the mechanisms by which food compounds regulate gene expression to modify cancer susceptibility. From a nutrition perspective, two key components potentially exert cancer chemopreventive effects: isothiocyanates (ITCs), present in cruciferous vegetables, and selenium (Se) which, as selenocysteine, is an integral part of selenoproteins. However, the role of these compounds in the expression of key selenoenzymes once the cancer process has been initiated still needs elucidation. Therefore, this investigation examined the effect of two forms of selenium, selenium-methylselenocysteine and sodium selenite, both individually and in combination with two ITCs, sulforaphane or iberin, on the expression of the two selenoenzymes, thioredoxin reductase 1 (TrxR1) and gastrointestinal glutathione peroxidase (GPx2), which are targets of ITCs, in Caco-2 cells. Co-treatment with both ITCs and Se induced expression of TrxR1 and GPx2 more than either compound alone. Moreover, pre-treatment of cells with ITC+Se enhanced cytoprotection against H(2)O(2)-induced cell death through a ROS-dependent mechanism. Furthermore, a single and double knockdown of TrxR1 and/or GPx2 suggested that both selenoproteins were responsible for protecting against H(2)O(2)-induced cell death. Together, these data shed new light on the mechanism of interactions between ITC and Se in which translational expression of the enhanced transcripts by the former is dependent on an adequate Se supply, resulting in a cooperative antioxidant protective effect against cell death.

UHRF1 regulation of the Keap1–Nrf2 pathway in pancreatic cancer contributes to oncogenesis

The cellular defence protein Nrf2 is a mediator of oncogenesis in pancreatic ductal adenocarcinoma (PDAC) and other cancers. However, the control of Nrf2 expression and activity in cancer is not fully understood. We previously reported the absence of Keap1, a pivotal regulator of Nrf2, in ∼70% of PDAC cases. Here we describe a novel mechanism whereby the epigenetic regulator UHRF1 suppresses Keap1 protein levels. UHRF1 expression was observed in 20% (5 of 25) of benign pancreatic ducts compared to 86% (114 of 132) of pancreatic tumours, and an inverse relationship between UHRF1 and Keap1 levels in PDAC tumours (n = 124) was apparent (p = 0.002). We also provide evidence that UHRF1‐mediated regulation of the Nrf2 pathway contributes to the aggressive behaviour of PDAC. Depletion of UHRF1 from PDAC cells decreased growth and enhanced apoptosis and cell cycle arrest. UHRF1 depletion also led to reduced levels of Nrf2‐regulated downstream proteins and was accompanied by heightened oxidative stress, in the form of lower glutathione levels and increased reactive oxygen species. Concomitant depletion of Keap1 and UHRF1 restored Nrf2 levels and reversed cell cycle arrest and the increase in reactive oxygen species. Mechanistically, depletion of UHRF1 reduced global and tumour suppressor promoter methylation in pancreatic cancer cell lines, and KEAP1 gene promoter methylation was reduced in one of three cell lines examined. Thus, methylation of the KEAP1 gene promoter may contribute to the suppression of Keap1 protein levels by UHRF1, although our data suggest that additional mechanisms need to be explored. Finally, we demonstrate that K‐Ras drives UHRF1 expression, establishing a novel link between this oncogene and Nrf2‐mediated cellular protection. Since UHRF1 over‐expression occurs in other cancers, its ability to regulate the Keap1–Nrf2 pathway may be critically important to the malignant behaviour of these cancers.

Understanding life and death decisions in human leukaemias.

Human leukaemia cells have an often unique ability to either undergo apoptotic cell death mechanisms or, at other times, undergo proliferative expansion, sometimes to the same stimulus such as the pluripotent cytokine TNFα (tumour necrosis factor α). This potential for life/death switching helps us to understand the molecular signalling machinery that underlies these cellular processes. Furthermore, looking at the involvement of these switching signalling pathways that may be aberrant in leukaemia informs us of their importance in cancer tumorigenesis and how they may be targeted pharmacologically to treat various types of human leukaemias. Furthermore, these important pathways may play a crucial role in acquired chemotherapy resistance and should be studied further to overcome in the clinic many drug-resistant forms of blood cancers. In the present article, we uncover the relationship that exists in human leukaemia life/death switching between the anti-apoptotic pro-inflammatory transcription factor NF-κB (nuclear factor κB) and the cytoprotective antioxidant-responsive transcription factor Nrf2 (nuclear factor-erythroid 2-related factor 2). We also discuss recent findings that reveal a major role for Btk (Brutons tyrosine kinase) in both lymphocytic and myeloid forms of human leukaemias and lymphomas.