Signal transducers and activators of transcription (STATs) are transcription factors activated in response to cytokines and growth factors. Constitutively active Stat3 has been shown to mediate oncogenic transformation in cultured cells and induce tumor formation in mice. An increasing number of tumor-derived cell lines as well as samples from human cancer have been reported to express constitutively active Stat3 protein. We previously demonstrated that ovarian cancer cell lines express high levels of constitutively active Stat3. In this study, we show that inhibition of the Stat3 signaling pathway using the Janus Kinase-selective inhibitor, AG490, and a dominant negative Stat3 (Stat3b) signi®cantly suppresses the growth of ovarian and breast cancer cell lines harboring constitutively active Stat3. In the ovarian cancer cell lines, AG490 also diminished the phosphorylation of Stat3, Stat3 DNA binding activity, and the expression of Bcl-x L . Further, AG490 induced signi®cant apoptosis in ovarian and breast cancer cell lines expressing high levels of constitutively active Stat3 but had a less profound e ect on normal cells lacking constitutively active Stat3. AG490 also enhanced apoptosis induced by cisplatin in ovarian cancer cells. These results suggest that inhibition of Stat3 signaling may provide a potential therapeutic approach for treating ovarian and breast cancers. Oncogene (2001) 20, 7925 ± 7934.

Angiotensin II (Ang II) is a peptide hormone that, like many cytokines, acts as a proinflammatory agent and growth factor. After injury to the liver, the hormone assists in tissue repair by stimulating hepatocytes and hepatic stellate cells to synthesize extracellular matrix proteins and secrete secondary cytokines and by stimulating myofibroblasts to proliferate. However, under conditions of chronic liver injury, all of these effects conspire to promote pathologic liver fibrosis. Much of this effect of Ang II results from activation of the proinflammatory NF-B transcription factor in response to stimulation of the type 1 Ang II receptor, a G protein-coupled receptor. Here, we characterize a previously undescribed signaling pathway mediating Ang II-dependent activation of NF-B, which is composed of three principal proteins, CARMA3, Bcl10, and MALT1. Blocking the function of any of these proteins, through the use of either dominant-negative mutants, RNAi, or gene targeting, effectively abolishes Ang II-dependent NF-B activation in hepatocytes. In addition, Bcl10 ؊/؊ mice show defective hepatic cytokine production after Ang II treatment. Evidence also is presented that this pathway activates NF-B through ubiquitination of IKK␥, the regulatory subunit of the I B kinase complex. These results elucidate a concrete series of molecular events that link ligand activation of the type 1 Ang II receptor to stimulation of the NF-B transcription factor. These findings also uncover a function of the CARMA, Bcl10, and MALT1 proteins in cells outside the immune system. G protein-coupled receptor ͉ hepatocyte ͉ IkB kinase ͉ inflammation ͉ ubiquitination

Abstract-The type-2 (AT 2 ) angiotensin (Ang) II receptor has been characterized as potentially counterregulatory to the actions of Ang II at its type-1 (AT 1 ) receptor. We investigated the effects of Ang II and CGP-42112A (CGP), a selective peptide AT 2 receptor agonist, on blood pressure (BP) in rats with or without pharmacological blockade of the AT 1 receptor with losartan (LOS) or valsartan (VAL). In anesthetized rats (nϭ5 per group) receiving normal sodium intake, Ang II (200 pmol/kg per minute IV) alone increased BP from a control of 112Ϯ3 to 168Ϯ7 mm Hg (PϽ0.001) and LOS (30 mg/kg) alone decreased BP to 89Ϯ7 mm Hg (PϽ0.0001 from control). Ang II administered together with LOS decreased BP further to 71Ϯ4 mm Hg (PϽ0.00001 from control and LOS alone). AT 2 receptor antagonist PD 123,319 (PD) completely blocked the hypotensive response to LOS combined with Ang II (PϭNS from control). In conscious rats (nϭ5 per group) receiving normal sodium intake, VAL (10 mg/kg) alone decreased BP from a control of 98Ϯ5 to 86Ϯ3 mm Hg (PϽ0.00001). Ang II combined with VAL induced a consistent, highly significant decline in BP for 6 days to a nadir of 69Ϯ3 mm Hg (PϽ0.01 versus daily VAL alone). PD completely blocked the chronic hypotensive response to the combination of Ang II and VAL to control levels before VAL administration. In another study in conscious rats (nϭ5 per group), CGP (70 g/kg per minute) also decreased BP in VAL-treated conscious rats. BP was 119Ϯ3 mm Hg during the control period, decreased to 86Ϯ6 mm Hg during 3 days of VAL alone, (PϽ0.00001) and decreased further to 65Ϯ7 mm Hg (PϽ0.001 from daily VAL alone) with 7 days of CGP in the presence of VAL. In the absence of VAL, CGP decreased BP for 4 consecutive days, and this response was blocked by PD. Also, the CGP-induced decrease in BP over a 7-day period was blocked by N G -nitro-L-arginine methyl ester, an inhibitor of NO synthase. The results strongly suggest that the AT 2 receptor induces a systemic vasodilator response mediated by NO that counterbalances the vasoconstrictor action of Ang II at the AT 1 receptor. (Hypertension. 2001;38:1272-1277.) Key Words: angiotensin II Ⅲ receptors, angiotensin II Ⅲ blood pressure Ⅲ losartan A ngiotensin (Ang) II is a plieotropic vasoactive peptide that acts at 2 known Ang II receptors, type 1 (AT 1 ) and type 2 (AT 2 ). 1 The actions of Ang II at AT 1 receptors are well characterized. However, the physiological actions of Ang II at AT 2 receptors have been difficult to elicit, at least in part because AT 2 receptors have a low degree of expression compared with that of AT 1 receptors. [1][2][3] Recent evidence suggests that AT 2 receptors may play a role in the regulation of arterial blood pressure (BP). 1,4 -15 In the mouse, targeted disruption of the AT 2 receptor gene increased BP slightly and induced pressor sensitivity to Ang II, both acutely and chronically. 4 -7 In the rat, the depressor phase of the biphasic BP response to Ang III (des-aspartyl[1]-Ang II) was blocked by AT 2 receptor blockade with PD 123,...

Proper regulation of nuclear factor-κB (NF-κB) transcriptional activity is required for normal lymphocyte function, and deregulated NF-κB signaling can facilitate lymphomagenesis. We demonstrate that the API2-MALT1 fusion oncoprotein created by the recurrent t(11;18)(q21;q21) in mucosa-associated lymphoid tissue (MALT) lymphoma induces proteolytic cleavage of NF-κB inducing kinase (NIK) at Arg325. NIK cleavage requires the concerted actions of both fusion partners and generates a C-terminal NIK fragment that retains kinase activity and is resistant to proteasomal degradation. The resulting deregulated NIK activity is associated with constitutive noncanonical NF-κB signaling, enhanced B-cell adhesion, and apoptosis resistance. Our study reveals the gain-of-function proteolytic activity of a fusion oncoprotein and highlights the importance of the noncanonical NF-κB pathway in B-lymphoproliferative disease.

Constitutive activation of the signal transducer and activator of transcription 3 (Stat3) and mutation of the p53 are both commonly detected in human prostate cancer cells. We sought to investigate whether there is functional regulation of Stat3 by wild-type (wt) p53. Our results demonstrate that expression of wt p53 but not mutant p53 signi®cantly reduced tyrosine phosphorylation of Stat3 and inhibited Stat3 DNA binding activity in both DU145 and Tsu prostate cancer cell lines that express constitutively active Stat3. Expression of the p53 downstream target, p21 WAF-1 , did not have any inhibitory e ect on Stat3 phosphorylation. Wt p53 but not p21 WAF-1 induced dramatic apoptosis in these prostate cancer cells. Expression of wt p53 did not cause a reduction of phosphorylation-independent Stat3 protein and reduction of phosphorylation of three unrelated protein kinases, ERK1, ERK2 (ERK1/2), and AKT. Interestingly, p53-dependent apoptosis occurred in the presence of high levels of phosphorylated AKT and ERK1/2 in both DU145 and Tsu prostate cancer cells. Further, we evaluated a series of established human prostate, breast, and ovarian cancer cell lines and found that all cancer cell lines expressing constitutively active Stat3, only harbor mutated or deleted p53. One implication of these results is that the anti-proliferative activities of p53 may not be compatible with the constitutive Stat3 signal in cancer cells.

The purpose of this study was to determine the precise role of angiotensin subtype-1 (AT1) and -2 (AT2) receptors and the mechanisms by which they act to alter fluid transport in the rat jejunum. In rats on normal sodium intake, ANG II at low dose stimulated net jejunal fluid absorption, whereas at a high dose the peptide inhibited absorption. Low-dose ANG II-stimulated fluid absorption was blocked completely by the specific AT2 receptor antagonist PD-123319 (PD) but was unchanged by the AT1receptor antagonist losartan (Los). The AT2 receptor agonist CGP-42112A, caused an inversely dose-dependent increase in fluid absorption, which also was totally prevented by PD but was unaltered by Los. Conversely, high-dose ANG II inhibition of absorption was blocked by Los but not by PD. In animals receiving normal sodium intake, neither Los nor PD alone altered fluid absorption. In sodium-restricted animals, however, Los alone increased absorption and PD alone inhibited absorption. In rats on normal sodium intake, low-dose ANG II increased jejunal interstitial and luminal (loop) fluid concentrations of cGMP. These increases in cGMP were blocked with PD but not with Los. 8-Bromoguanosine-3′,5′-cyclic monophosphate administered via the mesenteric artery or the submucosal interstitial space markedly increased absorption, but it inhibited absorption when administered into the loop. High-dose ANG II decreased jejunal interstitial and loop fluid cAMP and increased PGE2. The increase in PGE2 was blocked by Los but not by PD. The data demonstrate that ANG II mediates jejunal sodium and water absorption by an action at the AT2 receptor involving cGMP formation. The data also show that ANG II inhibits absorption via the AT1 receptor by a mechanism that is both negatively coupled to cAMP and increases jejunal PGE2 production.

The angiotensin (ANG) Type 2 (AT2) receptor is one of two major ANG II receptors that have been identified, cloned, and sequenced. Most of the biologic actions of ANG II are thought to be mediated by the AT1 receptor, but evidence is beginning to emerge that the AT2 receptor has a significant role in the regulation of blood pressure. In the adult rat, the AT2 receptor is expressed, albeit in low concentrations in kidney, mesenteric blood vessels, and heart. Most of the evidence suggests that the AT2 receptor stimulates a vasodilator signaling cascade that includes bradykinin, nitric oxide, and guanosine cyclic 3',5'-monophosphate. At lease some of the beneficial actions of AT1 receptor blockade are mediated by the AT2 receptor through this pathway. Several recent studies suggest that AT2 receptors may mediate vasodilation and hypotension. The AT2 receptor represents a potential therapeutic target for agonist action and a candidate molecule in the pathophysiology of hypertension.

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