Therapeutic potential of muscle growth promoters in a stress urinary incontinence model.

Weakness of urinary sphincter and pelvic floor muscles can cause insufficient urethral closure and lead to stress urinary incontinence. Bimagrumab is a novel myostatin inhibitor that blocks activin type II receptors, inducing skeletal muscle hypertrophy and attenuating muscle weakness. ß2-Adrenergic agonists, such as 5-hydroxybenzothiazolone derivative (5-HOB) and clenbuterol, can enhance muscle growth. We hypothesized that promoting muscle growth would increase leak point pressure (LPP) by facilitating muscle recovery in a dual-injury (DI) stress urinary incontinence model. Rats underwent pudendal nerve crush (PNC) followed by vaginal distension (VD). One week after injury, each rat began subcutaneous (0.3 mL/rat) treatment daily in a blinded fashion with either bimagrumab (DI + Bim), clenbuterol (DI + Clen), 5-HOB (DI + 5-HOB), or PBS (DI + PBS). Sham-injured rats underwent sham PNC + VD and received PBS (sham + PBS). After 2 wk of treatment, rats were anesthetized for LPP and external urethral sphincter electromyography recordings. Hindlimb skeletal muscles and pelvic floor muscles were dissected and stained. At the end of 2 wk of treatment, all three treatment groups had a significant increase in body weight and individual muscle weight compared with both sham-treated and sham-injured rats. LPP in DI + Bim rats was significantly higher than LPP of DI + PBS and DI + Clen rats. There were more consistent urethral striated muscle fibers, elastin fibers in the urethra, and pelvic muscle recovery in DI + Bim rats compared with DI + PBS rats. In conclusion, bimagrumab was the most effective for increasing urethral pressure and continence by promoting injured external urethral sphincter and pelvic floor muscle recovery.

Pharmacological Characterization of a Novel 5-Hydroxybenzothiazolone-Derived ß 2-Adrenoceptor Agonist with Functional Selectivity for Anabolic Effects on Skeletal Muscle Resulting in a Wider Cardiovascular Safety Window in Preclinical Studies.

The anabolic effects of ß 2-adrenoceptor (ß 2-AR) agonists on skeletal muscle have been demonstrated in various species. However, the clinical use of ß 2-AR agonists for skeletal muscle wasting conditions has been limited by their undesired cardiovascular effects. Here, we describe the preclinical pharmacological profile of a novel 5-hydroxybenzothiazolone (5-HOB) derived ß 2-AR agonist in comparison with formoterol as a representative ß 2-AR agonist that have been well characterized. In vitro, 5-HOB has nanomolar affinity for the human ß 2-AR and selectivity over the ß 1-AR and ß 3-AR. 5-HOB also shows potent agonistic activity at the ß 2-AR in primary skeletal muscle myotubes and induces hypertrophy of skeletal muscle myotubes. Compared with formoterol, 5-HOB demonstrates comparable full-agonist activity on cAMP production in skeletal muscle cells and skeletal muscle tissue-derived membranes. In contrast, a greatly reduced intrinsic activity was determined in cardiomyocytes and cell membranes prepared from the rat heart. In addition, 5-HOB shows weak effects on chronotropy, inotropy, and vascular relaxation compared with formoterol. In vivo, 5-HOB significantly increases hind limb muscle weight in rats with attenuated effects on heart weight and ejection fraction, unlike formoterol. Furthermore, changes in cardiovascular parameters after bolus subcutaneous treatment in rats and rhesus monkeys are significantly lower with 5-HOB compared with formoterol. In conclusion, the pharmacological profile of 5-HOB indicates superior tissue selectivity compared with the conventional ß 2-AR agonist formoterol in preclinical studies and supports the notion that such tissue-selective agonists should be investigated for the safe treatment of muscle-wasting conditions without cardiovascular limiting effects.

Mouse models of cancer-induced cachexia: Hind limb muscle mass and evoked force as readouts.

The majority of patients with advanced cancer suffer from cachexia, a systemic wasting syndrome, which subsequently impacts the tolerance to anti-cancer treatments, response to therapy, quality of life, and eventually, survival. Despite a high unmet medical need, there is currently no specific remedy available for an effective treatment of cachexia and its sequelae. A key feature of cachexia is the inexorable loss of skeletal muscle mass, which constitutes a main contributor to body weight loss and progressive functional impairments. Therefore, it's crucial to identify early readouts to detect and monitor the loss of muscle mass and function to initiate appropriate treatments timely. Here, we describe experimental cancer models using mouse (syngeneic) or human (xenograft) cancer cell lines with a rapid onset of tumor growth and cachexia. These models are easier to establish, monitor and reproduce compared to the genetically engineered mouse models currently available. Moreover, we establish readouts such as hind limb muscle mass and volume, as well as evoked force and food intake measurements, to allow the evaluation of potential therapeutic agents for the early treatment of cachexia and associated impairments.

Anti-tumor effect in human breast cancer by TAE226, a dual inhibitor for FAK and IGF-IR in vitro and in vivo.

Focal adhesion kinase (FAK) is a 125-kDa non-receptor type tyrosine kinase that localizes to focal adhesions. FAK overexpression is frequently found in invasive and metastatic cancers of the breast, colon, thyroid, and prostate, but its role in osteolytic metastasis is not well understood. In this study, we have analyzed anti-tumor effects of the novel FAK Tyr(397) inhibitor TAE226 against bone metastasis in breast cancer by using TAE226. Oral administration of TAE226 in mice significantly decreased bone metastasis and osteoclasts involved which were induced by MDA-MB-231 breast cancer cells and increased the survival rate of the mouse models of bone metastasis. TAE226 also suppressed the growth of subcutaneous tumors in vivo and the proliferation and migration of MDA-MB-231 cells in vitro. Significantly, TAE226 inhibited the osteoclast formation in murine pre-osteoclastic RAW264.7 cells, and actin ring and pit formation in mature osteoclasts. Moreover, TAE226 inhibited the receptor activator for nuclear factor κ B Ligand (RANKL) gene expression induced by parathyroid hormone-related protein (PTHrP) in bone stromal ST2 cells and blood free calcium concentration induced by PTHrP administration in vivo. These findings suggest that FAK was critically involved in osteolytic metastasis and activated in tumors, pre-osteoclasts, mature osteoclasts, and bone stromal cells and TAE226 can be effectively used for the treatment of cancer induced bone metastasis and other bone diseases.

Degenerative joint disease induced by repeated intra-articular injections of monosodium urate crystals in rats as investigated by translational imaging.

The objective of this work was to assess the consequences of repeated intra-articular injection of monosodium urate (MSU) crystals with inflammasome priming by lipopolysaccharide (LPS) in order to simulate recurrent bouts of gout in rats. Translational imaging was applied to simultaneously detect and quantify injury in different areas of the knee joint. MSU/LPS induced joint swelling, synovial membrane thickening, fibrosis of the infrapatellar fat pad, tidemark breaching, and cartilage invasion by inflammatory cells. A higher sensitivity to mechanical stimulus was detected in paws of limbs receiving MSU/LPS compared to saline-injected limbs. In MSU/LPS-challenged joints, magnetic resonance imaging (MRI) revealed increased synovial fluid volume in the posterior region of the joint, alterations in the infrapatellar fat pad reflecting a progressive decrease of fat volume and fibrosis formation, and a significant increase in the relaxation time T2 in femoral cartilage, consistent with a reduction of proteoglycan content. MRI also showed cyst formation in the tibia, femur remodeling, and T2 reductions in extensor muscles consistent with fibrosis development. Repeated intra-articular MSU/LPS injections in the rat knee joint induced pathology in multiple tissues and may be a useful means to investigate the relationship between urate crystal deposition and the development of degenerative joint disease.

Perampanel: a novel, orally active, noncompetitive AMPA-receptor antagonist that reduces seizure activity in rodent models of epilepsy.

PURPOSE: To assess the pharmacology of perampanel and its antiseizure activity in preclinical models. Perampanel [2-(2-oxo-1-phenyl-5-pyridin-2-yl-1,2-dihydropyridin-3-yl) benzonitrile] is a novel, orally active, prospective antiepileptic agent currently in development for refractory partial-onset seizures. METHODS: Perampanel pharmacology was assessed by examining changes in intracellular free Ca(2+) ion concentration ([Ca(2+) ](i) ) in primary rat cortical neurones, and [(3) H]perampanel binding to rat forebrain membranes. Antiseizure activity of orally administered perampanel was examined in amygdala-kindled rats and in mice exhibiting audiogenic, maximal electroshock (MES)-induced, pentylenetetrazole (PTZ) -induced, or 6 Hz-induced seizures. KEY FINDINGS: In cultured rat cortical neurones, perampanel inhibited α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-induced increases in [Ca(2+) ](i) (IC(50) 93 nm vs. 2 µm AMPA). Perampanel had a minimal effect on N-methyl-d-aspartate (NMDA)-induced increases in [Ca(2+) ](i) , and only at a high concentration (30 µm). [(3) H]Perampanel binding to rat forebrain membranes was not significantly displaced by glutamate or AMPA but was displaced by the noncompetitive AMPA receptor antagonists CP465022 (K(i) 11.2 ± 0.8 nm) and GYKI52466 (K(i) 12.4 ± 1 µm). In mice, perampanel showed protective effects against audiogenic, MES-induced, and PTZ-induced seizures (ED(50) s 0.47, 1.6, and 0.94 mg/kg, respectively). Perampanel also inhibited 6 Hz electroshock-induced seizures when administered alone or in combination with other antiepileptic drugs (AEDs). In amygdala-kindled rats, perampanel significantly increased afterdischarge threshold (p<0.05 vs. vehicle), and significantly reduced motor seizure duration, afterdischarge duration, and seizure severity recorded at 50% higher intensity than afterdischarge threshold current (p<0.05 for all measures vs. vehicle). Perampanel caused dose-dependent motor impairment in both mice (TD(50) 1.8 mg/kg) and rats (TD(50) 9.14 mg/kg), as determined by rotarod tests. In mice, the protective index (TD(50) in rotarod test/ED(50) in seizure test) was 1.1, 3.8, and 1.9 for MES-induced, audiogenic, and PTZ-induced seizures, respectively. In rat, dog, and monkey, perampanel had a half-life of 1.67, 5.34, and 7.55 h and bioavailability of 46.1%, 53.5%, and 74.5%, respectively. SIGNIFICANCE: These data suggest that perampanel is an orally active, noncompetitive, selective AMPA receptor antagonist with potential as a broad spectrum antiepileptic agent.

YM155, a novel survivin suppressant, enhances taxane-induced apoptosis and tumor regression in a human Calu 6 lung cancer xenograft model.

Survivin, an apoptotic inhibitor, is overexpressed in the majority of human tumor types and represents a novel target for anticancer therapy. Taxanes induce a mitotic cell-cycle block through the inhibition of microtubule depolymerization, with subsequent elevated expression/stabilization of survivin. We investigated the administration of survivin suppressant YM155 monobromide (YM155), in combination with docetaxel, in a human non-small-cell lung cancer (NSCLC) xenograft model. Animals received a 7-day continuous infusion of YM155, 2 mg/kg, and/or three bolus doses of docetaxel, 20 mg/kg, according to three dosing schedules: YM155 administered concomitantly with docetaxel, before docetaxel, and after docetaxel. YM155 administered either concomitantly with or before docetaxel showed significant antitumor activity (tumor regression ≥ 99%), with complete regression of the established human NSCLC-derived tumors in mice (eight of eight and seven of eight animals, respectively). Significantly fewer complete responses (three of eight animals) were achieved when YM155 was administered after docetaxel. No statistically significant decreases in body weight were observed in the combination versus docetaxel groups. YM155 administered concomitantly with docetaxel resulted in significant decreases in mitotic and proliferative indices, and in a significant increase in the apoptosis index. Elevated survivin expression was seen in tumors from mice treated with docetaxel alone; a significant reduction in survivin expression was seen in tumors from mice treated with YM155 alone or in combination with docetaxel, but not in the control group. These results indicate that in a human NSCLC xenograft model YM155 in combination with docetaxel diminished the accumulation of survivin by docetaxel and induced more intense apoptosis and enhanced antitumor activity, compared with single-agent YM155 or docetaxel.

Inhibition of podocyte FAK protects against proteinuria and foot process effacement.

Focal adhesion kinase (FAK) is a nonreceptor tyrosine kinase that plays a critical role in cell motility. Movement and retraction of podocyte foot processes, which accompany podocyte injury, suggest focal adhesion disassembly. To understand better the mechanisms by which podocyte foot process effacement leads to proteinuria and kidney failure, we studied the function of FAK in podocytes. In murine models, glomerular injury led to activation of podocyte FAK, followed by proteinuria and foot process effacement. Both podocyte-specific deletion of FAK and pharmacologic inactivation of FAK abrogated the proteinuria and foot process effacement induced by glomerular injury. In vitro, podocytes isolated from conditional FAK knockout mice demonstrated reduced spreading and migration; pharmacologic inactivation of FAK had similar effects on wild-type podocytes. In conclusion, FAK activation regulates podocyte foot process effacement, suggesting that pharmacologic inhibition of this signaling cascade may have therapeutic potential in the setting of glomerular injury.

Inhibition of focal adhesion kinase as a potential therapeutic strategy for imatinib-resistant gastrointestinal stromal tumor.

Focal adhesion kinase (FAK) is often up-regulated in a variety of malignancies, including gastrointestinal stromal tumor (GIST), and its overexpression seems to be associated with tumor progressiveness and poor prognosis. GIST is well known to have a mutation to c-KIT; thus, a specific c-KIT inhibitor (imatinib) is recognized as the first-line chemotherapy for GIST, although a certain type of c-KIT mutation reveals a resistance to imatinib due to as yet uncertain molecular mechanisms. To assess the c-KIT mutation-related variation of cellular responses to imatinib, murine lymphocyte-derived Ba/F3 cells, which are stably transduced with different types of c-KIT mutation, were treated with either imatinib or a FAK inhibitor (TAE226), and their antitumor effects were determined in vitro and in vivo. A mutation at exon 11 (KITdel559-560) displayed a high sensitivity to imatinib, whereas that at exon 17 (KIT820Tyr) showed a significant resistance to imatinib in vitro and in vivo. KIT820Tyr cells appeared to maintain the activities of FAK and AKT under the imatinib treatment, suggesting that FAK might play a role in cell survival in imatinib-resistant cells. When FAK activity in those cells was inhibited by TAE226, cell growth was equally suppressed and the cells underwent apoptosis regardless of the c-KIT mutation types. Oral administration of TAE226 significantly diminished tumor growth in nude mice bearing KIT(820Tyr) xenografts. In summary, c-KIT mutation at exon 17 displayed a resistance to imatinib with maintained activations of FAK and subsequent survival signals. Targeting FAK could be a potential therapeutic strategy for imatinib-resistant GISTs.

Autocrine activin A signalling in ovarian cancer cells regulates secretion of interleukin 6, autophagy, and cachexia.

BACKGROUND: The majority of patients with advanced cancer develop cachexia, a weight loss syndrome that severely reduces quality of life and limits survival. Our understanding of the underlying mechanisms that cause the condition is limited, and there are currently no treatment options that can completely reverse cachexia. Several tumour-derived factors and inflammatory mediators have been suggested to contribute to weight loss in cachectic patients. However, inconsistencies between studies are recurrent. Activin A and interleukin 6 (IL-6) are among the best studied factors that seem to be important, and several studies support their individual role in cachexia development. METHODS: We investigated the interplay between activin A and IL-6 in the cachexia-inducing TOV21G cell line, both in culture and in tumours in mice. We previously found that the human TOV21G cells secrete IL-6 that induces autophagy in reporter cells and cachexia in mice. Using this established cachexia cell model, we targeted autocrine activin A by genetic, chemical, and biological approaches. The secretion of IL-6 from the cancer cells was determined in both culture and tumour-bearing mice by a species-specific ELISA. Autophagy reporter cells were used to monitor the culture medium for autophagy-inducing activities, and muscle mass changes were evaluated in tumour-bearing mice. RESULTS: We show that activin A acts in an autocrine manner to promote the synthesis and secretion of IL-6 from cancer cells. By inhibiting activin A signalling, the production of IL-6 from the cancer cells is reduced by 40-50% (up to 42% reduction on protein level, P = 0.0048, and 48% reduction on mRNA level, P = 0.0308). Significantly reduced IL-6 secretion (P < 0.05) from the cancer cells is consistently observed when using biological, chemical, and genetic approaches to interfere with the autocrine activin A loop. Inhibiting activin signalling also reduces the ability of the cancer cells to accelerate autophagy in non-cancerous cells (up to 43% reduced autophagy flux, P = 0.0006). Coherent to the in vitro data, the use of an anti-activin receptor 2 antibody in cachectic tumour-bearing mice reduces serum levels of cancer cell-derived IL-6 by 62% (from 417 to 159 pg/mL, P = 0.03), and, importantly, it reverses cachexia and counteracts loss of all measured muscle groups (P < 0.0005). CONCLUSIONS: Our data support a functional link between activin A and IL-6 signalling pathways and indicate that interference with activin A-induced IL-6 secretion from the tumour has therapeutic potential for cancer-induced cachexia.

Dual tyrosine kinase inhibitor for focal adhesion kinase and insulin-like growth factor-I receptor exhibits anticancer effect in esophageal adenocarcinoma in vitro and in vivo.

PURPOSE: Focal adhesion kinase (FAK) regulates integrin and growth factor-mediated signaling pathways to enhance cell migration, proliferation, and survival, and its up-regulation correlates malignant grade and poor outcome in several types of cancer. In this study, we aimed to raise a potential therapeutic strategy using a FAK inhibitor for Barrett's esophageal adenocarcinoma. EXPERIMENTAL DESIGN: The expression status of FAK in clinical Barrett's esophageal adenocarcinoma tissues was determined by immunohistochemistry. Cultured esophageal adenocarcinoma cells were treated with TAE226, a specific FAK inhibitor with an additional effect of inhibiting insulin-like growth factor-I receptor (IGF-IR), to assess its anticancer effect in vitro. Western blot was carried out to explore a participating signaling pathway for TAE226-induced cell death. Furthermore, TAE226 was orally administered to s.c. xenograft animals to investigate its anticancer effect in vivo. RESULTS: Strong expression of FAK was found in 94.0% of Barrett's esophageal adenocarcinoma compared with 17.9% of Barrett's epithelia, suggesting that FAK might play a critical role in the progression of Barrett's esophageal adenocarcinoma. When esophageal adenocarcinoma cells were treated with TAE226, cell proliferation and migration were greatly inhibited with an apparent structural change of actin fiber and a loss of cell adhesion. The activities of FAK, IGF-IR, and AKT were suppressed by TAE226 and subsequent dephosphorylation of BAD at Ser(136) occurred, resulting in caspase-mediated apoptosis. In vivo tumor volume was significantly reduced by oral administration of TAE226. CONCLUSIONS: These results suggest that TAE226, a dual tyrosine kinase inhibitor for FAK and IGF-IR, could become a new remedy for Barrett's esophageal adenocarcinoma.

YM155, a novel small-molecule survivin suppressant, induces regression of established human hormone-refractory prostate tumor xenografts.

Various accumulating evidence suggests that survivin, a member of the inhibitor of apoptosis (IAP) family, plays an important role in drug resistance and cancer cell survival in many types of cancer, including hormone-refractory prostate cancer (HRPC). Here, we characterized YM155, a novel small-molecule survivin suppressant, using a survivin gene promoter activity assay. YM155 suppressed expression of survivin and induced apoptosis in PC-3 and PPC-1 human HRPC cell lines at 10 nmol/L. In contrast, YM155 up to 100 nmol/L showed little effect on expression levels of other IAP- or Bcl-2-related proteins. In a s.c. xenografted PC-3 tumor model in mice, 3-day continuous infusions of YM155 at 3 to 10 mg/kg induced massive tumor regression accompanied by suppression of intratumoral survivin. YM155 also completely inhibited the growth of orthotopically xenografted PC-3 tumors. No significant decreases in body weight were observed in mice treated with YM155 during the experimental period. Pharmacokinetic analyses indicated that YM155 is highly distributed to tumors and at concentrations approximately 20-fold higher than those in plasma. Our findings represent the first attempt to show tumor regression and suppression of survivin in p53-deficient human HRPC cells by a single small molecular compound treatment. Further extensive investigation of YM155 in many types of cancer, including HRPC, seems to be worthwhile to develop this novel therapeutic approach.

A novel tissue-selective ß2-adrenoceptor agonist with minimized cardiovascular effects, 5-HOB, attenuates neuropathic pain in mice.

OBJECTIVE: 5-HOB is a novel tissue selective, 5-hydroxybenzothiazolone-derived ß2 adrenoceptor agonist with minimized cardiovascular effects while retaining efficacy on skeletal muscle in preclinical experiments unlike conventional ß2 adrenoceptor agonists, however its effect on the nervous system has not been evaluated yet. Therefore, 5-HOB was evaluated in a mouse model of neuropathic pain. RESULTS: 5-HOB alleviated neuropathic allodynia in a dose dependent manner and reversed the changes in hind paw withdrawal thresholds to the sham control levels. The dose attenuating neuropathic allodynia was slightly lower than the dose inducing skeletal muscle hypertrophy. In conclusion, as reported with known ß2 adrenoceptor agonists, 5-HOB was also effective in attenuating neuropathic pain in mice in addition to its effect on skeletal muscle.

Evaluation of therapeutic effects of FAK inhibition in murine models of atherosclerosis.

OBJECTIVE: Therapeutic effects of focal adhesion kinase (FAK) inhibition using a small molecule inhibitor was evaluated in apolipoprotein E (apoE) knockout (KO) and low-density lipoprotein receptor (LDLr) KO mouse atherosclerosis models. RESULTS: The prevention trial consisted of an 8-week treatment with an FAK inhibitor concurrent treatment with a high fat (HF)/high cholesterol (HC) diet. The intervention trial consisted of 6- and 8-week treatment after 6- and 8-week pre-loading, respectively, of a HF/HC diet in apoE KO and LDLr KO mice, respectively. The inhibitor was admixed with a HF/HC diet and mice were given free access to the admixture. The FAK inhibitor exhibited marked inhibition against the development of the atherosclerosis in both of prevention and intervention trials at a dose of 0.03% without showing any remarkable toxic properties in biochemical examinations. These results indicated that FAK inhibition might be a possible candidate for novel therapeutic targets against atherosclerosis.

Identification and characterization of a phenyl-thiazolyl-benzoic acid derivative as a novel RAR/RXR agonist.

OBJECTIVE: To identify an agonist of RXRα and RARα with reduced undesired profiles of all-trans retinoic acid for differentiation-inducing therapy of acute promyelocytic leukemia (APL), such as its susceptibility to P450 enzyme, induction of P450 enzyme, increased sequestration by cellular retinoic acid binding protein and increased expression of P-glycoprotein, a virtual screening was performed. RESULTS AND CONCLUSION: In this study, a phenyl-thiazolyl-benzoic acid derivative (PTB) was identified as a potent agonist of RXRα and RARα. PTB was characterized in nuclear receptor binding, reporter gene, cell differentiation and cell growth assays. PTB bound directly to RXRα and RARα, but not to PPARα, δ(ß) or γ. PTB fully activated reporter genes with enhancer elements for RXRα/RXRα, and partially activated reporter genes with enhancer elements for RARα/RXRα, PPARδ(ß) and PPARγ. Furthermore, PTB induced differentiation and inhibited the growth of human APL cells. Thus, PTB is a novel dual agonist of RXRα and RARα and works as both a differentiation inducer and a proliferation inhibitor to leukemic cells.

Pharmacological profiling of a dual FAK/IGF-1R kinase inhibitor TAE226 in cellular and in vivo tumor models.

OBJECTIVE: A dual inhibitor of focal adhesion kinase (FAK) and insulin-like growth factor 1 receptor (IGF-1R), TAE226, was evaluated in a panel of cancer cell lines, MIA PaCa-2 human pancreatic tumor and 4T1 murine breast tumor models. The profiling data were generated during the drug discovery research prior to the first publication of TAE226 appeared in 2007 (Liu et al. in Mol Cancer Ther 6:1357-1367, 2007; Shi et al. in Mol Carcinog 46(6):488-496, 2007; Halder et al. in Cancer Res 67(22):10976-10983, 2007). RESULTS: In a panel of 37 cancer cell lines, TAE226 showed a mean GI50 value of 0.76 µmol/L. In the MIA PaCa-2 model, TAE226 inhibited phosphorylation of Y397-FAK and phosphorylation of S473-Akt as IGF-1R signaling in the cell culture in vitro and the tumor in mice. Oral administration of TAE226 induced tumor stasis at 30 mg/kg and tumor regression at 100 mg/kg in the subcutaneous tumor, and inhibited the orthotopic tumor growth in a dose-dependent manner. Similarly in the 4T1 model, TAE226 inhibited phosphorylation of Y397-FAK and S473-Akt in the cell culture in vitro and the tumor in mice. Oral administration of TAE226 inhibited the orthotopic tumor growth and metastasis to the lung in a dose-dependent manner. Thus, TAE226 represents a novel class of selective and small molecule kinase inhibitor with a potent in vivo activity.

Group IVA phospholipase A2 is associated with the storage of lipids in adipose tissue and liver.

Prostaglandin (PG) E(2) is considered to participate in the storage of fat in adipocytes and hepatocytes, but roles of group IVA phospholipase A(2) (PLA(2)), a key PLA(2) isozyme in the arachidonic acid cascade, remain unclear. The present study examined the possible involvement of the enzyme using group IVA PLA(2)-deficient mice (C57BL/6 background, 22 weeks of age) fed a normal diet (5.3% fat). The ratio of epididymal fat pad weight to body weight was significantly reduced in group IVA PLA(2)-deficient mice compared to wild-type mice. Histological analysis revealed that in group IVA PLA(2)-deficient mice, the adipocytes were smaller, and hepatocytes bearing cytoplasmic vacuolation were scarce. Hepatic triglyceride content and the serum levels of PGE(2) in the deficient mice were also lower. However, there was no difference in the serum levels of insulin, glucose, non-esterified free fatty acid, or total cholesterol between the deficient and wild-type mice. Our findings suggest that group IVA PLA(2) is involved in the storage of lipids in the adipose tissue and liver and in determining circulating PGE(2) levels.

TAE226, a dual inhibitor for FAK and IGF-IR, has inhibitory effects on mTOR signaling in esophageal cancer cells.

Esophageal cancer is one of the most aggressive cancers in the world. Novel preventive and therapeutic strategies tend to target the key molecules involved in the signaling transduction pathways for cell growth. It is known that FAK and mTOR are important controllers of cell growth. TAE226, a novel small molecule compound, is a potent ATP competitive inhibitor of FAK and IGF-IR. TAE226 can block FAK and IGF-IR signaling pathways. The purpose of this study was to explore the inhibitory effects on mTOR signaling and the mechanism of cell growth suppression by TAE226. We examined the expression of mTOR and S6 in esophageal cancer cells (SEG-1) and normal esophageal epithelial cells (KOB-13) and the efficacy of TAE226 against SEG-1 cells. mTOR and S6 were overexpressed in SEG-1 cells compared with KOB-13 cells. TAE226 inhibited the expression of mTOR, Akt, p70S6K and S6 as well as the phosphorylation of mTOR (Ser2448), Akt (Ser473), p70S6K (Thr389) and S6 (Ser240/244). As a result, TAE226 induced a dose-dependent decrease in cell growth (number) and damage in the cell shape. Together, these data show that TAE226 has potent inhibitory effects on mTOR signaling and esophageal cancer cell growth indicating that TAE226 has potential application in esophageal cancer treatment.

Inhibition of both focal adhesion kinase and insulin-like growth factor-I receptor kinase suppresses glioma proliferation in vitro and in vivo.

Multiple genetic aberrations in human gliomas contribute to their highly infiltrative and rapid growth characteristics. Focal adhesion kinase (FAK) regulates tumor migration and invasion. Insulin-like growth factor-I receptor (IGF-IR), whose expression correlates with tumor grade, is involved in proliferation and survival. We hypothesized that inhibiting the phosphorylation of FAK and IGF-IR by NVP-TAE226 (hereafter called TAE226), a novel dual tyrosine kinase inhibitor of FAK and IGF-IR, would suppress the growth and invasion of glioma cells. In culture, TAE226 inhibited extracellular matrix-induced autophosphorylation of FAK (Tyr(397)). TAE226 also inhibited IGF-I-induced phosphorylation of IGF-IR and activity of its downstream target genes such as MAPK and Akt. TAE226 retarded tumor cell growth as assessed by a cell viability assay and attenuated G(2)-M cell cycle progression associated with a decrease in cyclin B1 and phosphorylated cdc2 (Tyr(15)) protein expression. TAE226 treatment inhibited tumor cell invasion by at least 50% compared with the control in an in vitro Matrigel invasion assay. Interestingly, TAE226 treatment of tumor cells containing wild-type p53 mainly exhibited G(2)-M arrest, whereas tumor cells bearing mutant p53 underwent apoptosis. Induction of apoptosis by TAE226 was substantiated by detection of caspase-3/7 activation and poly(ADP-ribose) polymerase cleavage and by an Annexin V apoptosis assay. More importantly, TAE226 treatment significantly increased the survival rate of animals in an intracranial glioma xenograft model. Collectively, these data show that blocking the signaling pathways of FAK and IGF-IR with TAE226 has the potential to be an efficacious treatment for human gliomas.

Epigenetic targeting of bromodomain protein BRD4 counteracts cancer cachexia and prolongs survival.

Cancer cachexia is a devastating metabolic syndrome characterized by systemic inflammation and massive muscle and adipose tissue wasting. Although it is responsible for approximately one-third of cancer deaths, no effective therapies are available and the underlying mechanisms have not been fully elucidated. We previously identified the bromodomain and extra-terminal domain (BET) protein BRD4 as an epigenetic regulator of muscle mass. Here we show that the pan-BET inhibitor (+)-JQ1 protects tumor-bearing mice from body weight loss and muscle and adipose tissue wasting. Remarkably, in C26-tumor-bearing mice (+)-JQ1 administration dramatically prolongs survival, without directly affecting tumor growth. By ChIP-seq and ChIP analyses, we unveil that BET proteins directly promote the muscle atrophy program during cachexia. In addition, BET proteins are required to coordinate an IL6-dependent AMPK nuclear signaling pathway converging on FoxO3 transcription factor. Overall, these findings indicate that BET proteins may represent a promising therapeutic target in the management of cancer cachexia.