Dexamethasone (Dex) and radiation therapy are established modalities in multiple myeloma. In this study, we propose a novel combination of Dex plus radiation that shows superior clonogenic cell killing and apoptosis of myeloma cells and selectively eliminates myeloma cells when cocultured with bone marrow stromal cells (BMSCs). Dex was found to inhibit the release of interleukin-6 from irradiated BMSCs, which is an established myeloma cell proproliferative cytokine. In 5TGM1 model, the combination of Dex with skeletal targeted radiotherapy (153-Sm-EDTMP) prolonged median survival time and inhibited radiation-induced myelosuppression. A two-cycle treatment of Dex plus 153-Sm-EDTMP was well tolerated and further improved median survival time. Mechanistically, Dex increased superoxide and hydrogen peroxide production and augmented radiation-induced oxidative stress and cell death of myeloma cells. In contrast, Dex inhibited radiation-induced increase in pro-oxidant levels and enhanced the clonogenic survival in normal hematopoietic stem and progenitor cells. Treatment with either N-acetylcysteine or the combination of polyethylene glycol (PEG)-conjugated copper, zinc-superoxide dismutase, and PEG-catalase significantly protected myeloma cells from Dex-induced clonogenic death. Overall, these results demonstrate that Dex in combination with radiotherapy enhances the killing of myeloma cells while protecting normal bone marrow hematopoiesis through a mechanism that involves selective increases in oxidative stress.

Regulator of G protein signaling 6 (RGS6) is a member of a family of proteins called RGS proteins, which function as GTPase-activating proteins (GAPs) for G␣ subunits. Given the role of RGS6 as a G protein GAP, the link between G protein activation and cancer, and a reduction of cancer risk in humans expressing a RGS6 SNP leading to its increased translation, we hypothesized that RGS6 might function to inhibit growth of cancer cells. Here, we show a marked down-regulation of RGS6 in human mammary ductal epithelial cells that correlates with the progression of their transformation. RGS6 exhibited impressive antiproliferative actions in breast cancer cells, including inhibition of cell growth and colony formation and induction of cell cycle arrest and apoptosis by mechanisms independent of p53. RGS6 activated the intrinsic pathway of apoptosis involving regulation of Bax/Bcl-2, mitochondrial outer membrane permeabilization (MOMP), cytochrome c release, activation of caspases-3 and -9, and poly(ADP-ribose) polymerase cleavage. RGS6 promoted loss of mitochondrial membrane potential (⌬⌿ m ) and increases in reactive oxygen species (ROS). RGS6-induced caspase activation and loss of ⌬⌿ m was mediated by ROS, suggesting an amplification loop in which ROS provided a feed forward signal to induce MOMP, caspase activation, and cell death. Loss of RGS6 in mouse embryonic fibroblasts dramatically impaired doxorubicin-induced growth suppression and apoptosis. Surprisingly, RGS6-induced apoptosis in both breast cancer cells and mouse embryonic fibroblasts does not require its GAP activity toward G proteins. This work demonstrates a novel signaling action of RGS6 in cell death pathways and identifies it as a possible therapeutic target for treatment of breast cancer. Regulator of G protein signaling (RGS)2 proteins comprise a family of proteins, defined by the presence of a semiconserved region called the RGS domain, which negatively regulate heterotrimeric G protein signaling (1-3). These proteins were discovered as essential negative regulators of heterotrimeric G protein signaling by genetic studies in yeast followed by similar studies in Caenorhabditis elegans (4 -6). RGS proteins function as GTPase-activating proteins (GAPs) for heterotrimeric G protein subunits (7,8), an activity bestowed by their RGS domain (9) thereby enhancing the shut-off mechanism for G protein signaling. Thirty mammalian genes encode proteins with the hallmark RGS domain or less closely related versions of this domain, and RGS proteins have been classified into subfamilies based upon sequence similarities within the RGS domain that extend to sequences outside of this domain (2, 8). RGS6 is a member of the R7 subfamily of proteins that possess, in addition to their RGS domain, a GGL (G ␥ subunit-like) domain that allows for association with G␤ 5 and stabilization of RGS6 and a Drosophila/EGL10/Pleckstrin homology/DEP helical extension (DEP/DHEX) domain that allows for association with one of two additional proteins, R7BP or R9AP, and is thought to pr...

IL (interleukin)-6, an established growth factor for multiple myeloma cells, induces myeloma therapy resistance, but the resistance mechanisms remain unclear. The present study determines the role of IL-6 in re-establishing intracellular redox homoeostasis in the context of myeloma therapy. IL-6 treatment increased myeloma cell resistance to agents that induce oxidative stress, including IR (ionizing radiation) and Dex (dexamethasone). Relative to IR alone, myeloma cells treated with IL-6 plus IR demonstrated reduced annexin/propidium iodide staining, caspase 3 activation, PARP [poly(ADP-ribose) polymerase] cleavage and mitochondrial membrane depolarization with increased clonogenic survival. IL-6 combined with IR or Dex increased early intracellular pro-oxidant levels that were causally related to activation of NF-κB (nuclear factor κB) as determined by the ability of N-acetylcysteine to suppress both pro-oxidant levels and NF-κB activation. In myeloma cells, upon combination with hydrogen peroxide treatment, relative to TNF (tumour necrosis factor)-α, IL-6 induced an early perturbation in reduced glutathione level and increased NF-κB-dependent MnSOD (manganese superoxide dismutase) expression. Furthermore, knockdown of MnSOD suppressed the IL-6-induced myeloma cell resistance to radiation. MitoSOX Red staining showed that IL-6 treatment attenuated late mitochondrial oxidant production in irradiated myeloma cells. The present study provides evidence that increases in MnSOD expression mediate IL-6-induced resistance to Dex and radiation in myeloma cells. The results of the present study indicate that inhibition of antioxidant pathways could enhance myeloma cell responses to radiotherapy and/or chemotherapy.

Gallic acid and gallate esters are widely used as dietary supplements or additives with clinical significances. Over the last few decades, a large number of publications have been reported stating the antioxidative, antiapoptotic, cardioprotective, neuroprotective, and anticancer properties of gallic acid and gallates, and mostly demonstrated their antioxidative or prooxidative properties influencing the reactive oxygen species (ROS) signaling networks. However, very little focus has been paid to clinical trials, and this restricted their use as a prescribed preventative supplement. Since mitochondria are the principal organelles responsible for ROS generation, we reviewed the existing literature of mitochondria-specific effects of gallates including ROS production, respiration, mitochondrial biogenesis, apoptosis, and the physico-chemical parameters affecting the outcome of gallate supplementation to various health scenarios such as cardiovascular diseases, neurodegeneration, hepatic ailments, or cancers. The major signaling pathways and the molecules targeted by gallic acid and its derivatives have also been discussed with emphasis on mitochondria as the target site. This review provides a better understanding of the effect of gallic acid and gallate esters on mitochondrial functions and in designing effective preventative measures against the onset of various diseases.

The creatine/creatine kinase (CK) system plays a key role in cellular energy buffering and transport. In vertebrates, CK has four isoforms expressed in a tissue-specific manner. In the process of creatine biosynthesis several other important metabolites are formed. The anticancer effect of creatine had been reported in the past, and recent literature has reported low creatine content in several types of malignant cells. Furthermore, creatine can protect cardiac mitochondria from the deleterious effects of some anticancer compounds. Previous work from our laboratory showed progressive decrease of phosphocreatine, creatine and CK upon transformation of skeletal muscle into sarcoma. It was convincingly demonstrated that prominent expression of creatine-synthesizing enzymes L-arginine: glycine amidinotransferase and N-guanidinoacetate methyltransferase occurs in sarcoma, Ehrlich ascites carcinoma and sarcoma 180 cells; whereas, both these enzymes are virtually undetectable in skeletal muscle. Creatine transporter also remained unaltered in malignant cells. The anticancer effect of methylglyoxal had been known for a long time. The present work shows that this anticancer effect of methylglyoxal is significantly augmented in presence of creatine. On creatine supplementation the effect of methylglyoxal plus ascorbic acid was further augmented and there was no visible sign of tumor. Moreover, creatine and CK, which were very low in sarcoma tissue, were significantly elevated with the concomitant regression of tumor.