Oxidative Stress Plays a Critical Role in Inactivating Mutant BRAF by Geldanamycin Derivatives

Department of Radiation Oncology, Washington University School of Medicine, St. Louis, Missouri.
Cancer Research (Impact Factor: 9.33). 09/2008; 68(15):6324-30. DOI: 10.1158/0008-5472.CAN-07-6602
Source: PubMed


The geldanamycin derivatives 17-allylamino-17-demethoxygeldanamycin (17-AAG) and 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) are promising chemotherapeutic drugs that inhibit heat shock protein 90 (HSP90) function. Previous studies have shown that 17-AAG/DMAG treatment induces the degradation of mutant BRAF (V600E) and inhibits the activation of mitogen-activated protein/extracellular signal-regulated kinase 1/2 (MEK1/2). We have found, however, that HSP90 inhibition alone is not sufficient for efficient BRAF(V600E) degradation in some cells. HSP90 inhibitors structurally unrelated to geldanamycin, radicicol and novobiocin, while inducing the degradation of the HSP90 client protein RAF-1 fail to induce BRAF(V600E) degradation or inhibit MEK1/2 activation in HT29 human colon cancer cells. Moreover, after treatment with 17-DMAG, the kinase activity of residual, undegraded BRAF(V600E) was also lost. Incubation of cells with a reactive oxygen species (ROS) scavenger, N-acetyl cysteine, partially restored kinase activity and also partially prevented BRAF(V600E) degradation due to 17-DMAG treatment. Conversely, treatment with the ROS producing drug menadione clearly inhibited MEK1/2 and reduced BRAF(V600E). These results suggest that in addition to direct inhibition of HSP90, the antitumor effect of geldanamycin and its derivatives is also mediated though the production of ROS, which may directly inactivate tumorigenic mutant BRAF(V600E).

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    • "Interestingly, not only expression of TGFb1 is increased under the state of increased OS but also ROS can be downstream intermediates of TGFb1 (Poli et al. 2004). On the other hand, one study showed that under in vitro conditions, certain ROS-enhancing agents could somehow paradoxically induce degradation of the BRAF mutant (Fukuyo et al. 2008). Thus, BRAF mutant, TGFb1, and ROS may have a complex relationship and role in the pathogenesis of thyroid cancer. "
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    ABSTRACT: Oxidative stress (OS) is a state of excessive free radicals and reactive metabolites among which the most important class is reactive oxygen species (ROS) - radicals derived from oxygen - as represented by the superoxide anion radical (O2(·-)) and its reactive metabolites, hydroxyl radical (·OH) and hydrogen peroxide (H(2)O(2)). In essence, OS represents an imbalance between the production of oxidants - ROS - and their elimination by antioxidative systems in the body. Many studies have linked OS to thyroid cancer by showing its association with abnormally regulated oxidative or antioxidative molecules. The study by Wang et al. in the December 2011 issue of Endocrine-Related Cancer (18, 773-782) further supports this relationship by demonstrating a high total oxidant status and OS index in thyroid cancer patients. The origin of ROS in thyroid cancer patients has not been defined, but thyroid cancer itself can be one since inflammation, a major event in it, is a classical source of ROS. ROS may in turn enhance the mitogen-activated protein (MAP) kinase and phosphatidylinositol-3-kinase (PI3K) pathways, forming a vicious cycle propelling thyroid tumorigenesis. Regardless of the mechanism, the clinical implication of the association of OS with thyroid cancer is severalfold: one, OS is a new risk factor for thyroid cancer; two, OS confers thyroid cancer patients an increased risk for cardiovascular diseases, degenerative neurological disorders, and other cancers that are classically associated with OS; and three, interference with OS may reduce this risk and be therapeutically beneficial to thyroid cancer itself in thyroid cancer patients. These interesting possibilities deserve further studies.
    Endocrine Related Cancer 12/2011; 19(1):C7-11. DOI:10.1530/ERC-11-0360 · 4.81 Impact Factor
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    • "Recent investigations have suggested that oxidative stress caused by inflammation may be in part responsible for mutations of the BRAF gene (Martinez- Cadenas et al. 2011). Other authors also found that oxidative stress plays a critical role in inactivating mutant BRAF by geldanamycin derivatives (Fukuyo et al. 2008). These findings are similar to those of this study. "
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    ABSTRACT: Oxidative stress is considered to be involved in the pathophysiology of all cancers. In order to evaluate the total oxidant/antioxidant status in patients with thyroid cancer and to investigate the relationship between oxidative stress parameters and serum thyroid profiles among thyroid cancer patients and various controls, we determined oxidative status including total antioxidant status (TAS) and total oxidant status (TOS) and calculation of oxidative stress index (OSI) in sera in 82 thyroid cancer patients, 56 benign thyroid disease patients, and 50 healthy controls. It was found that serum TAS levels were significantly lower in patients with thyroid cancer than in controls (P<0.001), while serum TOS levels and OSI values were significantly higher (both P<0.001) in the cancer patients. No significant correlations were observed between various oxidative stress markers and thyroid profiles in either the thyroid cancer patients or the controls. Receiver operating characteristic curve analysis demonstrated that OSI was the best indicator for distinguishing cancer patients from benign thyroid diseased or healthy controls, followed by TOS and TAS. Risk estimate statistics also indicated that TOS and/or OSI were good risk factors to discriminate patients with thyroid cancer from two controls. These findings suggested that oxidants are increased and antioxidants are decreased in patients with thyroid cancer. OSI may be a more useful oxidative stress biomarker than TAS and TOS for monitoring the clinical status of thyroid cancer patients.
    Endocrine Related Cancer 12/2011; 18(6):773-82. DOI:10.1530/ERC-11-0230 · 4.81 Impact Factor
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    • "These findings suggest that Raf and MEK inhibitors could be combined with cytostatic drugs or radiation in the therapy of cancer. The relationships between Raf and oxidative stress are manifold: RAF/MEK/ERK activation can prevent the onset of oxidative stress in growth factor-deprived cells (Kuznetsov et al., 2008); on the other hand, generation of reactive oxygen species by derivatives of geldanamycin, a chemotherapeutic that inhibits the chaperone function of HSP90 and enforces the degradation of their client proteins, including Raf, is able to inhibit the activity of BRAFV600E (Fukuyo et al., 2008). Finally, a most interesting connection, relevant in terms of oncogene-selective therapy, has been reported recently between oncogenic activation of RAS and RAF, and the small-molecule drug erastin. "
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    ABSTRACT: Raf are conserved, ubiquitous serine/protein kinases discovered as the cellular elements hijacked by transforming retroviruses. The three mammalian RAF proteins (A, B and CRAF) can be activated by the human oncogene RAS, downstream from which they exert both kinase-dependent and kinase-independent, tumor-promoting functions. The kinase-dependent functions are mediated chiefly by the MEK/ERK pathway, whose activation is associated with proliferation in a broad range of human tumors. Almost 10 years ago, activating BRAF mutations were discovered in a subset of human tumors, and in the past year treatment with small-molecule RAF inhibitors has yielded unprecedented response rates in melanoma patients. Thus, Raf qualifies as an excellent molecular target for anticancer therapy. This review focuses on the role of BRAF and CRAF in different aspects of carcinogenesis, on the success of molecular therapies targeting Raf and the challenges they present.
    Oncogene 05/2011; 30(32):3477-88. DOI:10.1038/onc.2011.160 · 8.46 Impact Factor
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