Abstract
The aberrant activation of STAT3 occurs in many human cancers and promotes tumor progression. Phosphorylation of a tyrosine at amino acid Y705 is essential for the function of STAT3. Synthesized carbazole derived with fluorophore compound 12 was discovered to target STAT3 phosphorylation. Compound 12 was found to inhibit STAT3-mediated transcription as well as to reduce IL-6 induced STAT3 phosphorylation in cancer cell lines expressing both elevated and low levels of phospho-STAT3 (Y705). Compound 12 potently induced apoptosis in a broad number of TNBC cancer cell lines in vitro and was effective at inhibiting the in vivo growth of human TNBC xenograft tumors (SUM149) without any observed toxicity. Compound 12 also effectively inhibited the growth of human lung tumor xenografts (A549) harboring aberrantly active STAT3. In vitro and in vivo studies showed that the inhibitory effects of 12 on phospho-STAT3 were through up-regulation of the protein–tyrosine phosphatase PTPN6. Our present studies strongly support the continued preclinical evaluation of compound 12 as a potential chemotherapeutic agent for TNBC and cancers with constitutive STAT3 signaling.
Introduction
Current emphasis in cancer drug development has shifted from nonspecific cytotoxic chemotherapies to molecularly targeted reagents.(1) The etiology of cancer is multimodal, but great progress has been made in recent times toward the understanding of these diseases. Discovery of oncogenes(2) and transcription factors(3, 4) that play the critical roles in regulating gene expression with the development of cancers has aided in the development of new approaches for the cancer treatment. Signal transducers and activators of transcription 3 (STAT 3),(5-7) one of seven known members of the mammalian STAT family, regulates transcription of several genes upon activation of membrane receptors by various cytokines.(8) Ligand binding to cytokine receptors triggers activation of cytokine receptor associated kinases such as Janus kinases (JAKs) or receptor tyrosine kinases.(8) Activation of STAT3 is achieved by phosphorylation of Y705 residue by these cytokine receptor associated kinases.(8) Phosphorylated STAT3 translocates into the nucleus to bind STAT3 responsive elements and regulates expression of target genes.(7) Phosphorylation of STAT3 is also regulated by protein tyrosine phosphatases such as tyrosine–protein phosphatase nonreceptor type 6 (PTPN6/SHP-1), tyrosine–protein phosphatase nonreceptor type 11 (PTPN11/SHP-2), and T-cell protein–tyrosine phosphatase (TC-PTP).(9)
Constitutive STAT3 activation has been detected in a large number of human cancers, and elevated levels of the activated STAT3 are associated with a poor clinical prognosis.(10, 11) Studies showed that the inhibition of STAT3 expression attenuated the proliferation and survival of STAT3-positive cancer cell lines in vitro and in vivo, while normal tissues were unaffected by loss of STAT3.(5, 12-14) Furthermore, the causal role of STAT3 in oncogenesis has been confirmed via molecular biology and pharmacology tools in disease-relevant models.(15) All of these support STAT3 as a molecular target for cancer drug discovery,(9, 15-17) however, only one STAT3 inhibitor has been advanced into clinical trials(18) because of the physicochemical problems of the studied molecules.(16) To address those problems that are an obstacle to the STAT3 inhibitors’ being promoted into clinical trials, further studies have been carried out in the field recently and many advances have been achieved as the results.(12, 19-22)
A wide range of synthetic analogues based on natural products have emerged as therapeutically useful antitumor agents.(23) Among these natural products, natural and synthetic congeners of the carbazole alkaloids comprise a large group of therapeutically useful antitumor agents in the various stages of clinical development.(24-26) Although earlier reports(27, 28) have demonstrated that some of carbazoles negatively regulate the STAT3-mediated transcription and/or STAT3-DNA binding and phospho-STAT3 with relatively high concentration (30–50 μM), the mechanisms of these regulations still remain unknown. We have been interested in targeting STAT3 pathways.(29, 30) In an effort to discover targeted therapeutics to treat cancers, we have developed fluorescent small molecules, which could be observed directly as it interacts with receptor-positive cell lines, as potential anticancer agents.(31-33) Meanwhile, our previously studies(34) showed that the antiproliferative effect on the cancer cell lines of the free carbazole is much less than the corresponding scaffolds with dansyl group. Herein, novel imageable carbazoles (Figure 1) were designed and synthesized, and their biological activities as STAT3 inhibitors were further evaluated.
Figure 1. Novel carbazoles fluorophore.
Results
Chemistry
A palladium-catalyzed cross-coupling reaction(35) was carried out (Scheme 1) between the commercially available compounds tert-butyl 4-iodo-2-methylphenylcarbamate (1) and 4-methoxy-2-nitrophenylboronic acid (2), providing the intermediate tert-butyl 4′-methoxy-3-methyl-2′-nitrobiphenyl-4-ylcarbamate (3) in a yield of 79%. Deprotection of 3 in the presence of 20% trifluoroacetic acid–dichloromethane and followed by dansylation with dansyl chloride resulted in the formation of the fluorescent compound 4. Treatment of 4 with triphenylphosphine in 1,2-dichlorobenzene under reflux(36, 37) overnight resulted in the reductive cyclization of the 2-nitrobiphenyl to form a mixture of two carbazole 5 and 6 (ca. 1:1), which were readily separable by chromatography. The structural assignment of the two isomers was based on the 1H NMR coupling patterns of the aryl protons. The 1H NMR spectrum of compound 6 showed two doublets (δ 6.48 and 7.46, J = 8.4 Hz) corresponding to the protons on carbons 3 and 4, respectively, whereas the 1H NMR spectrum of compound 5 showed two singlets (δ 6.61 and 7.52) corresponding to the protons on carbons 1 and 4.
Scheme 1. Synthesis of Compounds 5 and 6a
a Reagents and conditions: (a) Pd(PPh3)4, K2CO3, CH3CN–H2O, reflux, 79%; (b) (i) 20% TFA–DCM, (ii) dansyl chloride, pyridine, 80%; (c) PPh3, 1,2-DCB, reflux under N2, 42% for 5 and 38% for 6.
The known compound 11(34) was prepared in an improved way as shown in Scheme 2. The commercially available reagents 7 and 8 were coupled under the standard Suzuki reaction condition, providing the biphenyl-4-ol compound 9 (80%). After dansylation (10) and reductive cyclization, two isomers, 7-methoxy-1-methyl-9H-carbazol-2-yl 5-(dimethylamino)naphthalene-1-sulfonate (11) and 7-methoxy-3-methyl-9H-carbazol-2-yl 5-(dimethylamino)naphthalene-1-sulfonate,(34) were formed. Because our previously studies indicated that 11 is around 20 times stronger than the corresponding isomer to the cancer cell lines,(34) several conditions were investigated to increase the ratio of 11 to its isomer. We found that the removal of oxygen atmosphere could increase the yield of 11 from 40% to 60% in the presence of triphenylphosphine and 1,2-dichlorobenzene under reflux. The demethylation was carried out in the presence of BBr3 in dichloromethane. Concerned that the dansyl ester is good leaving group, the deprotection was performed at low temperature (−78 °C) and the process of the demethylation was monitored by TLC. The result of TLC indicated that compound 11 was intact at low temperature (−78 °C to −30 °C), even up to 8 h. When the mixture was warmed up to ambient temperature, the demethylation proceeded cleanly and afforded the phenol product 12 with a yield of 81%. Compound 12 was treated with dansyl chloride in anhydrous dichloromethane and triethylamine, resulting in the didansylated compound 13.
Scheme 2. Synthesis of Compounds 12 and 13a
aReagents and conditions: (a) Pd(PPh3)4, K2CO3, CH3CN–H2O, reflux, 80%; (b) dansyl chloride, DCM–TEA, 85%; (c) PPh3, 1,2-DCB, reflux under N2, 60%; (d) BBr3, dichloromethane, −78 °C to room temperature, 81%; (e) dansyl chloride, DCM–TEA, 71%.
Biology
Compound 12 Inhibits the STAT3-Mediated Transcription and IL-6-Induced Phosphorylation of STAT3 in Triple-Negative Breast Cancer (TNBC) Cells
To determine the biological effect of our novel compounds and a natural product carbazole mahanine(38, 39) on the STAT3 pathway, we performed a typical reporter gene assay. HS578T cells were transfected with the expression vector for a constitutively active STAT3 (STAT3C)(40) and the APRE-Luc reporter (a STAT3-responsive reporter).(41) The cells were treated with 1 μM of compounds for 24 h, and the luciferase reporter activity was measured (Figure 2A). From this screening, compound 12 was identified as the most potent analogue in reduce the STAT3C-induced reporter gene activity in HS578T cells at 1 μM concentration. The effect of compound 12 on STAT3C-mediated transcription was comparable to that of AZD1480,(42) a JAK2 kinase inhibitor (Figure 2A). In HS578T cells transfected with STAT3C expression vector and the APRE-Luc, 12 repressed the STAT3C-dependent transactivation in a dose-dependent manner (Figure 2B). The effect of 12 on the STAT3 pathway was also assessed in the interleukin-6 (IL-6) induction model. HS578T and MCF7 cells, transfected with the APRE-Luc, were pretreated for 1 h with an increasing amount of compound 12 followed by 2 h (for HS578T) or 16 h (for MCF-7) activation with 5 ng/mL of IL-6. Under these conditions, compound 12 significantly reduced the IL-6-induced activation of endogenous STAT3-mediated transcription (Figure 2C). In addition, 12 reduced the IL-6-induced phosphorylation of STAT3 (Y705) in a dose-dependent manner in two separate TNBC cell lines, HS578T and SUM149PT (Figure 2D).
Figure 2. Repression of STAT3C-mediated transcription by compound 12. (A) Reporter gene assay for STAT3C transactivation in HS578T cells. HS578T cells were transfected with the APRE-Luc and STAT3C expression vector and further treated with 1 μM of compounds for 24 h. (B) HS578T cells were transfected with the APRE-Luc and STAT3C expression vector and further treated with increasing amount of compound 12 for 24 h. (C) HS578T and MCF7 cells were transfected with the APRE-Luc and treated with increasing amount of compound 12 for 1 h. The cells were further treated with 5 ng/mL of IL-6 for 2 h for HS578T or 16 h for MCF7 before reporter gene assay. (A–C) The relative reporter activities are presented as mean ± SD from experiments performed in triplicate. *P < 0.05, **P < 0.01, and ***P < 0.001. (D) Compound 12 inhibits the IL-6-induced phosphorylation of STAT3. HS578T cells were treated with 12 for 2 h as indicated. The IL-6 (5 ng/mL) was added to the culture medium and incubated for 30 min. The cell lysates were analyzed by the indicated antibodies and β-actin was used as a loading control.
Compound 12 Exhibits the Antiproliferative Effects on a Broad Range of Cancer Cell Lines
Because activation of STAT3 has been implicated in human cancers, we determined the level of phospho-STAT3 (Y705) in a panel of breast cancer cell lines by Western blot analysis. Interestingly, the level of phospho-STAT3 (Y705) was elevated in all the TNBC cell lines tested as compared to two luminal breast cancer cells, T47D and MCF7 (Figure 3A). We further determined the cytotoxic effects of 12 on the panel of TNBC cell lines. The cells were treated with increasing concentration of 12 for ∼72 h, and the viable cells were measured by MTT assay. Interestingly, compound 12 demonstrated antiproliferative affects on a broad range of TNBC cell lines (Figure 3B). Compound 12 also exhibited the antiproliferative effects on additional cancer cell lines, including A431 (a squamous carcinoma cell line), A549 (a lung cancer cell line) and PC-3 (a prostate cancer cell line) with GI50 values 0.16 μM (for A431), 2.5 μM (for A549), and 3 and 7.9 μM (for PC-3), respectively, in MTT assay with ∼72 h treatment (data not shown).
Figure 3. Antiproliferative effects and induction of PTPN6 by 12 in TNBC cells. (A) The elevated expression of phospho-STAT3 (Y705) in TNBC cell lines. The cell lysates from exponentially growing cells were analyzed by indicated antibodies. β-Actin was used as a loading control. (B) EC50 values of 12 were determined by MTT cell viability assay. Data from two independent experiments performed in triplicate are shown as mean ± SEM (C) HS578T and SUM149PT cells were treated with increasing amount of 12 for 24 h and the cell lysates were analyzed by indicated antibodies and β-actin was used as a loading control. (D) RT-PCR analysis of cells treated as in (C), GAPDH was used as a loading control. (E). The cells were treated with compound 12 or CP690550 in the absence or presence of increasing amount of pervanadate for 16 h. The cell lysates were analyzed by indicated antibodies, and β-actin was used as a loading control. (F) The cells were transfected with either control- or PTPN6-siRNA for 3 days and further treated with 12 for 24 h. The cell lysates were subjected to Western blot analysis with indicated antibodies.
Compound 12 Reduces Phospho-STAT3 by Inducing Protein–Tyrosine Phosphatase PTPN6
Because elevated phospho-STAT3 was detected in various TNBC cell lines, we further treated two TNBC cell lines with compound 12 without extracellular IL-6 stimulation and the level of phospho-STAT3 was determined by Western blot analysis. As shown in Figure 3C, 12 reduced the phospho-STAT3 (Y705) in a dose-dependent manner. We also found that the level of cyclin D1, a transcriptional target of STAT3, was also reduced by compound 12 treatment.
The effect of compound 12 on the mRNA expressions of STAT3 targets was assessed by semiquantitative reverse-transcription PCR. Consistent with Western blot analysis, the level of cyclin D1 mRNA was decreased by compound 12 in two TNBC cells (Figure 3D). Under these conditions, the mRNA expression of survivin, another STAT3 target was also reduced by compound 12. Interestingly the mRNA expression of PTPN6 was induced by compound 12 in a dose-dependent manner (Figure 3D).
To determine the effect of PTPN6 upregulation on STAT3, HS578T, and SUM149PT, cells were treated with compound 12 or CP690550 (a JAK1/2 inhibitor)(43) in the presence of a phosphatase inhibitor, pervanadate, for 16 h, and the level of phospho-STAT3 was determined by Western blot analysis. Consistent with the induction of PTPN6 mRNA, the compound 12 mediated reduction of phospho-STAT3 (Y705) was reversed by increasing the amount of pervanadate (Figure 3E). On the contrary, the kinase inhibitor CP690550-mediated inhibition of phospho-STAT3 was not affected by pervanadate.
To further verify the PTPN-dependent dephosphorylation of phospho-STAT3, we performed siRNA-based knockdown of PTPN6 in HS578T cells. HS578T cells were transfected with either control- or PTPN6-siRNA for 3 days and further treated with compound 12 for 24 h, and then the level of proteins were determined by Western blot analysis. Consistent with PTPN6 mRNA expression, treatment of compound 12 induced PTPN6 protein expression in HS578T cells transfected with control-siRNA (Figure 3F). As expected, transfection of PTPN6-siRNA reduced the compound 12 mediated PTPN6 protein expression. Under these conditions, knockdown of PTPN6 reversed the dephosphorylation of STAT3. Taken together, these results suggest that compound 12 inhibits phospho-STAT3 (Y705) by induction of PTPN6 phosphatase in TNBC cells.
Imaging of Compound 12 in Human Breast Cancer Cells
Compound 12 contains a dansyl moiety, which allows it to visibly fluoresce at 552 nm when excited at 370 nm (Figure 4). After treatment of the human breast cancer MDA-MB-231 cells with compound 12, a multiphoton laser was used to excite the compound, and its emission from within the cells was observed by two-photon confocal microscopy (Figure 4). Compound 12 (shown in green) was present within the cytoplasm of the cells but not in the nuclei (Figure 4a).
Figure 4. Compound 12 fluoresces in the cytoplasm of MDA-MB-231 cells. MDA-MB-231 cells were treated with 10 μM of compound 12 for 1 h. The fluorophore of compound 12 was excited at 725 nm with a multiphoton laser and imaged with a 500–550 nm filter. (a) Compound 12 fluorescence shown in green. (b) Differential interference contrast (DIC) image. (c) Parts a and b are merged.
In Vivo Studies Indicated 12 Significantly Reduces Nonsmall-Cell (A549) Lung Tumor and Human Breast Cancer (SUM 149) Growth in Xenograft Mice
Athymic Balb/c nude mice bearing A549 human nonsmall cell lung tumor that harbor aberrantly active STAT3(44-46) were dosed via intraperitoneal injection with 10 mg/kg compound 12 at every other day for 27 days (total 13 doses, N = 5, Figure 5). The onset of the antitumor growth effect of compound 12 occurred on day 5 (inhibition rate 68.7%) with the maximum inhibitory effect occurring on day 9 (inhibition rate 73.76%). The duration of inhibition lasted up to 27 days (P < 0.05), with tumor growth inhibition rate between 54.94 to 73.76% during this period.
Figure 5. Treatment with compound 12 reduces the volume of human nonsmall cell (A549) lung xenograft tumors. Mice bearing A549 tumors were administered 12 via ip 10 mg/kg every other day. Tumor sizes were measured every other day and converted into volumes by L × w × w/2, plotted against days of treatment. Data are present as the mean ± SEM (n = 5, P < 0.05).
Athymic Balb/c nude mice bearing human TNBC tumor (SUM149) were dosed via intraperitoneal injection with 10 and 30 mg/kg compound 12 at every other day for 5 days (Figure 6A). No significant changes in body weight or obvious signs of toxicity such as loss of appetite, decreased activity, or lethargy were observed during the efficacy study.
Figure 6. Treatment with compound 12 reduces the volume of human breast cancer (SUM149) xenograft tumors. (A) Mice bearing SUM149 tumors were administered compound 12 via ip 10 and 30 mg/kg every other day. Tumor sizes were measured every other day and converted into volumes by L × w × w/2, plotted against days of treatment. Data are present as the mean ± SEM (n = 5, P < 0.05). (B) Western blot of day 5 for compound 12 treated SUM149 tumors were evaluated for PTPN6, STAT3, and pSTAT3. GAPDH was used as a control. (C) Densitometry of PTPN6 with statistical significance using a one tailed t test (* P < 0.005).
Compound 12 was effective at directly inhibiting the growth of the SUM149 xenograft and resulted in a 57% reduction in tumor volume at day 5 (Figure 6A). Western analysis of the SUM149 tumors treated with compound 12 also revealed an up-regulation of PTPN6 (statistical significant at 30 mg/kg, Figure 6B) and pSTAT3 (Figure 6C). This data strongly supports the potential for use and further preclinical examination of compound 12 in TNBC.
Acute toxicity was evaluated by using the acute toxicity up-and-down procedure,(47) where animals were dosed with compound 12 via ip injection and observed for 14 days. The estimated LD50 of 550 mg/kg was determined, with the 95% PL confidence interval between 116.4 and 811 mg/kg.
Discussion and Conclusions
Conventional chemotherapeutic agents do not distinguish between healthy and cancer cells. As a newer type of cancer treatment, targeted therapy uses drugs to more precisely identify and attack cancer cells while doing little damage to normal cells. The aberrant activation of STAT3 occurs in the majority of human cancers and its cancer promoting activities have been confirmed involves with cell proliferation, metastasis, angiogenesis, host immune evasion, and resistance to apoptosis.(48, 49) Given its importance to cancer, STAT3 is considered a molecular target for the development of new cancer therapy.
Phosphorylation of a tyrosine at amino acid Y705 is essential for the function of STAT3, and elevated levels of STAT3 phosphorylation have been correlated with tumor invasion, nodal metastasis, and staging.(50, 51) Phospho-STAT3 has gained much attention,(52-54) and although several phospho-STAT3 inhibitors have been reported,(18, 55-57) only one of them have reached the clinic for several reasons including lacking of specificity and scaffold suitability. Herein we described the discovery and characterization of a novel carbazole compound 12 that strongly inhibits STAT3-mediated transcription as well STAT3 phosphorylation induced by IL-6. IL-6 is the most notable activator of STAT3 pathway and crucial for inducing and maintaining a cancer-promoting inflammatory environment, tumor cell survival through the up-regulation of antiapoptotic genes, and metastasis of tumor cells.(5) Recently, it has been demonstrated that TNBC cell lines have autocrine secretion of high levels of inflammatory cytokines including IL-6, IL-8, and CXCL1.(58) Consistent with this, we found that TNBC cell lines expressed a high level of phospho-STAT3 without exogenous IL-6 and compound 12 also reduced the activated phospho-STAT3 in these cell lines as well (Figure 7).
Figure 7. STAT3 pathway (A) and mechanism of compound 12’s STAT3 pathway inhibition (B).
The observed cytoplasmic fluorescence of compound 12 provides visual evidence that the compound enters human breast cancer cells. Further, it provides strong support for molecular target engagement of compound 12 in the cytoplasm and provides a new tool for study of STAT3 mechanistic pathways involved in mediating protein up-regulation via cytoplasmic signaling.
We found that the inhibition of phospho-STAT3 by compound 12 was via the up-regulation of cytoplasmic protein–tyrosine phosphatase PTPN6. Although loss of PTPN6 has been shown to enhance JAK3/STAT3 signaling in ALK-positive anaplastic large-cell lymphoma,(59) the roles of PTPN6 in TNBC are not yet understood. One emerging question in our study relates to how compound 12 induces the expression of PTPN6. Because the loss of PTPN6 expression is due to its promoter DNA methylation that is mediated by STAT3/DNMT1,(60) we are currently investigating the potential role of compound 12 in the epigenetic regulation of gene expression in TNBC cells via a cytoplasmic control.
Compound 12 was effective at inhibiting the growth of both human TNBC xenograft tumors (SUM149) and human xenograft (A549) tumors that harbor aberrantly active STAT3 without any observed toxicity. Our present studies strongly support the continued preclinical evaluation of compound 12 as a potential chemotherapeutic agent for TNBC and cancers with constitutive STAT3 signaling.
Experimental Section
All reagents and solvents were purchased from commercial suppliers and used as received unless noted otherwise. Flash column chromatography separations were done on a Biotage SP1 system monitoring at 254 and 310 nm. NMR spectra were recorded on a Varian 400 spectrometer at 25 °C, operating at 400 MHz for 1H and 100 MHz for 13C NMR. The chemical shifts are expressed in ppm downfield from TMS as an internal standard. Reactions were monitored by thin-layer chromatography (TLC) on silica gel 60 glass slides. The structure of the synthesized compounds follows unequivocally from the mode of synthesis and the m/z values found in their low- and high-resolution mass spectra, TLC, and NMR spectroscopy verified the purity. The HPLC system from Shimadzu consisted of a CBM-20A connector, LC-20AD pumps, an SDP-10AV UV detector, and a SIL-HTA autosampler. Compounds 12 and 13 were monitored at 254 nm by the UV detector. A Symmetry C18 column with 4.6 mm × 250 mm and 5 μm from Waters was used. The mobile phase for 12 consisted of 70% acetonitrile and 30% water containing 0.1% acetic acid or 80% acetonitrile and 20% water containing 0.1% acetic acid. The mobile phase for 13 consisted of 70% acetonitrile and 30% water containing 0.1% acetic acid or 90% acetonitrile and 10% water containing 0.1% acetic acid. The flow rate of mobile phase was 1.0 mL/min. The retention time of 12 was 9.0 or 4.7 min and 13 was 6.4 or 5.1 min. HPLC showed that the purity of both of the compounds is ≥95%.
tert-Butyl 4′-Methoxy-3-methyl-2′-nitrobiphenyl-4-ylcarbamate (3)
4-Methoxy-2-nitrophenylboronic acid 2 (1.07 g, 5.5 mmol) was added to a solution of tert-butyl 4-iodo-2-methylphenylcarbamate 1 (1.66 g, 5.0 mmol) in CH3CN–H2O (20 mL, v/v = 2:1), followed by Pd(PPh3)4 (115 mg, 0.1 mmol) and K2CO3 (2.65 g, 25 mmol). The mixture was refluxed for 16 h. After cooling to room temperature, the mixture was filtered through a Celite pad and the filtrate was concentrated to remove acetonitrile. The residue was diluted with DCM (100 mL) and washed with saturated brine (2 × 30 mL), and the organic phase was dried over Na2SO4 and concentrated. tert-Butyl 4′-methoxy-3-methyl-2′-nitrobiphenyl-4-ylcarbamate (3) (1.42 g, 79%) was afforded after concentration and chromatography (hexane/ethyl acetate = 6:1); mp 136 °C (from ethanol). 1H NMR (400 MHz, CDCl3), δ 7.89 (d, J = 8.4 Hz, 1 H), 7.33–7.29 (m, 2 H), 7.12 (d, J = 2.4 Hz, 1 H), 7.10 (d, J = 2.8 Hz, 1 H), 7.06 (brs, 1 H), 6.31 (brs, 1 H), 3.88 (s, 3 H), 2.26 (s, 3 H), 1.53 (s, 9 H). 13C NMR (100 MHz, CDCl3), δ 158.8, 1559.9, 152.8, 149.5, 136.2, 132.7, 132.1, 129.8, 128.2, 126.5, 120.5, 118.5, 108.9, 80.6, 55.8, 28.3 (3 C), 17.7. Anal. Calcd for C19H22N2O5: C 63.67, H 6.19, N, 7.82. Found: C 63.59, H 6.16, N 7.77.
5-(Dimethylamino)-N-(4′-methoxy-3-methyl-2′-nitrobiphenyl-4-yl)naphthalene-1-sulfonamide (4)
A solution of 3 (1.0 g, 2.79 mmol) in 20% trifluoroacetic acid–dichloromethane (15 mL) was stirred at room temperature for 5 h and concentrated to dryness. The residue was added to EtOH (2 × 3 mL) and concentrated to completely remove the residual trifluoroacetic acid. The product was used for next step without further purification. Dansyl chloride (2.25 g, 8.37 mmol) was added to a solution of the fresh prepared residue in pyridine (15 mL) at 0 °C and warmed up to room temperature. After stirring for 24 h, the mixture was concentrated and chromatographed (hexane/ethyl acetate = 3:1) to afford the 5-(dimethylamino)-N-(4′-methoxy-3-methyl-2′-nitrobiphenyl-4-yl)naphthalene sulfonamide (4) (1.1 g, 80% over two steps). 1H NMR (400 MHz, CDCl3), δ 8.55 (d, J = 8.4 Hz, 1 H), 8.35 (d, J = 8.8 Hz, 1 H), 8.20 (dd, J = 1.2 Hz, 7.6 Hz, 1 H), 7.57 (dd, J = 8.0 Hz, 8.8 Hz, 1 H), 7.48 (t, J = 8.4 Hz, 1 H), 7.30 (d, J = 2.4 Hz, 1 H), 7.20 (m, 2 H), 7.12 (d, J = 9.2 Hz, 1 H), 7.08 (dd, J = 3.2 Hz, 8.4 Hz, 1 H), 6.93 (m, 2 H), 6.54 (s, 1 H), 3.86 (s, 3 H), 2.89 (s, 6 H), 2.00 (s, 3H). 13C NMR (100 MHz, CDCl3), 159.0, 149.4, 134.7, 134.4, 132.6, 131.0, 130.8, 130.2, 129.5, 128.4, 127.6, 126.5, 123.2, 118.6, 108.9, 55.8, 45.4, 17.6. HRMS (TOF-ES+): m/z [M + 1]+ calcd for C26H25N3O5S, 492.1593; found, 492.1660.
5-(Dimethylamino)-N-(7-methoxy-3-methyl-9H-carbazol-2-yl)naphthalene-1-sulfonamide (5) and 5-(Dimethylamino)-N-(7-methoxy-1-methyl-9H-carbazol-2-yl)naphthalene-1-sulfonamide (6)
Triphenylphosphine (1.32 g, 5.0 mmol) was added to a solution of 5 (980 mg, 2 mmol) in 1,2-dichlorobenzene (10 mL). The mixture was refluxed under N2 for 48 h, concentrated, and chromatographed (hexane–ethyl acetate–methanol 4:1:0.01) to afford 5-(dimethylamino)-N-(7-methoxy-3-methyl-9H-carbazol-2-yl)naphthalene-1 sulfonamide (5) (390 mg, 42%) and 5-(dimethylamino)-N-(7-methoxy-1-methyl-9H-carbazol-2-yl)naphthalene-1-sulfonamide (6) (350 mg, 38%).
5: 1H NMR (400 MHz, CDCl3), δ 8.50 (d, J = 8.4 Hz, 1 H), 8.41 (d, J = 8.4 Hz, 1 H), 8.12 (dd, J = 1.2 Hz, 7.2 Hz, 1 H), 7.82 (s, 1 H), 7.74 (d, J = 8.4 Hz, 1 H), 7.53 (dd, J = 7.6 Hz, 8.8 Hz, 1 H), 7.52 (s, 1 H), 7.38 (dd, J = 7.6 Hz, 8.4 Hz, 1 H), 7.18 (d, J = 7.2 Hz, 1 H), 6.82 (d, J = 2.0 Hz, 1 H), 6.78 (dd, J = 7.0 Hz, 8.8 Hz, 1 H), 6.61 (s, 1 H), 3.86 (s, 3 H), 2.88 (s, 6 H), 2.01 (s, 3 H). 13C NMR (100 MHz, CDCl3), δ 159.0, 141.3, 138.2, 134.8, 131.2, 130.5, 130.2, 129.6, 128.3, 123.2, 122.5, 121.5, 120.7, 120.6, 116.4, 115.2, 108.2, 106.1, 94.6, 55.6, 45.4, 17.8. HRMS (TOF-ES+): m/z [M + 1]+ calcd for C26H26N3O3S, 460.1695; found, 460.1720; Anal. Calcd for C26H25N3O3S + 1.2H2O: C 64.90, H 5.74, N, 8.73. Found: C 64.83, H 5.49, N 8.45.
6: 1H NMR (400 MHz, CDCl3), δ 8.52 (d, J = 8.4 Hz, 1 H), 8.41 (d, J = 8.4 Hz, 1 H), 8.00 (dd, J = 1,2 Hz, 7.2 Hz, 1 H), 7.81 (s, 1 H), 7.78 (d, J = 8.8 Hz, 1 H), 7.56 (dd, J = 8.0 Hz, 8.8 Hz, 1 H), 7.46 (d, J = 8.4 Hz, 1 H), 7.37 (dd, J = 7.6 Hz, 8.4 Hz, 1 H), 7.20 (d, J = 7.6 Hz, 1 H), 6.89 (d, J = 2.0 Hz, 1 H), 6.80 (dd, J = 7.0 Hz, 8.8 Hz, 1 H), 6.48 (t, 4.0 Hz, 2 H), 3.87 (s, 3 H), 2.90 (s, 6 H), 2.28 (s, 3 H). 13C NMR (100 MHz, CDCl3), δ 159.1, 155.2, 141.2, 139.1, 134.5, 131.2, 130.5, 130.4, 129.9, 129.7, 128.4, 123.1, 121.1, 118.8, 117.2, 116.8, 115.1, 108.5, 94.7, 55.6, 45.4, 12.1. HRMS (TOF-ES+): m/z [M + 1]+ calcd for C26H26N3O3S, 460.1695; found, 460.1719. Crystallization from ethyl acetate: mp 165–168 °C. Anal. Calcd for C26H25N3O3S + 0.25EtOAc: C 67.34, H 5.65, N, 8.73. Found: C 67.06, H 5.71, N 8.63.
4′-Methoxy-3-methyl-2′-nitrobiphenyl-4-ol (9)
4-Hydroxyphenylboronic acid (8) (1.5 g, 10 mmol) was added to a solution of 4-iodo-3-nitroanisole (7) (3.1 g, 11 mmol) in CH3CN–H2O (30 mL, v/v = 2:1), followed by Pd(PPh3)4 (350 mg, 0.3 mmol) and K2CO3 (5.3 g, 50 mmol), the mixture was refluxed overnight. After cooling to room temperature, the mixture was filtered through a Celite pad. The filtrate was concentrated, diluted with DCM (100 mL), and washed with saturated brine (3 × 30 mL). After concentration and chromatography (hexane/acetone = 6:1) afforded 4′-methoxy-3-methyl-2′-nitrobiphenyl-4-ol (9) (2.1 g, 80%). 1H NMR (400 MHz, CDCl3), δ 7.31 (m, 2 H), 7.12 (dd, J = 2.8 Hz, 8.4 Hz, 1 H), 7.04 (s, 1 H), 6.98 (d, J = 8.0 Hz, 1 H), 6.78 (d, J = 8.0 Hz, 1 H), 3.88 (s, 3 H), 2.26 (s, 3 H). 13C NMR (100 MHz, CDCl3), δ 158.7, 153.7, 132.7, 130.6, 129.5, 128.3, 126.7, 124.1, 118.5, 115.1, 108.8, 55.8, 15.7. MS (TOF-ES+): 260.3 m/z [M + 1]+.
4′-Methoxy-3-methyl-2′-nitrobiphenyl-4-yl 5-(Dimethylamino)naphthalene-1-sulfonate (10)
Dansyl chloride (3.14 g, 12 mmol) was added to a solution of 9 (1.5 g, 5.8 mmol) in DCM–TEA (15 mL, v/v, 3:1) with stirring at 0 °C. The mixture was allowed to room temperature and stirred overnight, diluted with DCM (90 mL), and washed with saturated brine (30 mL). After drying over Na2SO4 and concentration, the residue was purified via chromatography (hexane/EtOAc = 5:1) to afforded 4′-methoxy-3-methyl-2′-nitrobiphenyl-4-yl 5-(dimethylamino)naphthalene-1-sulfonate (10) (2.41 g, 85%). 1H NMR (400 MHz, CDCl3), δ 8.67 (d, J = 8.8 Hz, 1 H), 8.49 (d, J = 8.8 Hz, 1 H), 8.19 (dd, J = 1.2 Hz, 7.2 Hz, 1 H), 7.65 (dd, J = 7.6 Hz, 8.8 Hz, 1 H), 7.53 (dd, J = 7.6 Hz, 8.4 Hz, 1 H), 7.33 (d, J = 2.4 Hz, 1 H), 7.25 (m, 2 H), 7.10 (dd, J = 2.8 Hz, 8.8 Hz, 1 H), 7.08 (d, J = 2.4 Hz, 1 H), 6.86 (dd, J = 2.0 Hz, 8.4 Hz, 1 H), 6.66 (d, J = 8.4 Hz, 1 H), 3.87 (s, 3 H), 2.93 (s, 6 H), 2.21 (s, 3 H). 13C NMR (100 MHz, CDCl3), δ 159.2, 149.4, 148.3, 135.9, 132.6, 132.1, 132.0, 131.8, 131.2, 130.8, 130.0, 128.9, 127.4, 126.5, 123.1, 121.8, 118.6, 115.7, 109.0, 55.9, 45.4, 16.6. MS (TOF-ES+): 493.5 m/z [M + 1]+. Anal. Calcd for C26H24N2O6S: C 63.40, H 4.91, N, 5.69. Found: C 63.79, H 4.59, N 5.45.
7-Methoxy-1-methyl-9H-carbazol-2-yl 5-(dimethylamino)naphthalene-1-sulfonate (11)
A solution of compound 10 (200 mg, 0.41 mmol) and triphenylphosphine (534 mg, 2.03 mmol) in 1,2-DCB (10 mL) was vacuumed and flushed with N2 for three times. The mixture was refluxed for 15 h and concentrated. After chromatography (hexane/ethyl acetate = 5;1) gave 7-methoxy-1-methyl-9H-carbazol-2-yl 5-(dimethylamino)naphthalene-1-sulfonate (11) (117 mg, 60%). NMR data were identical with those of reported.(34)
7-Hydroxy-1-methyl-9H-carbazol-2-yl 5-(dimethylamino)naphthalene-1-sulfonate (12)
1.0 M solution of BBr3 (0.22 mL, 0.22 mmol) was added to the solution of compound 11 (48 mg, 0.11 mmol) in anhydrous dichloromethane (5 mL) under N2 with stirring at −78 °C dropwisely. The mixture was warmed up to ambient temperature within 1 h and stirred overnight. The reaction was cooled with ice–water and diluted with dichloromethane (40 mL), and saturated aq NaHCO3 (3.0 mL) was added within 15 min. After of the addition, the mixture was washed with saturated brine aq (20 mL) and the organic phase was dried over Na2SO4 and concentrated, then chromatographed (toluene/acetone = 6:1) to afford 12 (37 mg, 81%, purity >97% based on HPLC analysis, see Supporting Information). 1H NMR (400 MHz, acetone-d6), δ 10.17 (s, 1 H), 8.71 (dt, J = 1.2 Hz, 8.8 Hz, 1H), 8.54 (dt, J = 0.8 Hz, 8.8 Hz, 1 H), 8.38 (brs, 1 H), 8.11(dd, J = 1.2 Hz, 7.2 Hz, 1 H), 7.79 (d, J = 8.4 Hz, 1 H), 7.45 (dd, J = 7.6 Hz, 8.8 Hz, 1 H), 7.61 (dd, J = 7.6 Hz, 8.4 Hz, 1 H), 7.55 (d, J = 8.8 Hz, 1H), 7.36 (dd, J = 0.8 Hz, 7.6 Hz, 1 H), 6.93 (dd, J = 0.4 Hz, 2.0 Hz, 1 H), 6.73 (dd, J = 2.4 Hz, 8.4 Hz, 1 H), 6.34 (d, J = 8.4 Hz, 1 H), 2.92 (s, 6 H), 2.37 (s, 3 H). 13C NMR (100 MHz, acetone-d6), δ 156.7, 152.1, 145.1, 142.4, 139.7, 132.2, 131.8, 130.8, 130.0, 129.8, 128.9, 123.2, 121.7, 120.8, 119.3, 116.3, 115.9, 115.6, 113.7, 112.7, 109.0, 96.7, 44.7, 10.6. HRMS (TOF-ES+): m/z [M + 1]+ calcd for C25H22N2O4S, 447.1379; found, 447.1399. Crystallized from toluene: mp 99–103 °C. Anal. Calcd for C25H22N2O4S +0.25toluene: C 68.42, H 5.15, N, 5.97. Found: C 68.17, H 5.32, N 5.89.
1-Methyl-9H-carbazole-2,7-diyl Bis(5-(dimethylamino)naphthalene-1-sulfonate) (13)
Dansyl chloride was added to a solution of 12 (30 mg, 0.067 mmol) in anhydrous dichloromethane (2 mL) in the presence of Et3N (0.5 mL) under N2 with stirring at 0 °C. After stirring for 2 h, the mixture was concentrated and chromatographed (hexane/acetone = 3:1) to give compound 13 (32 mg, 71%, purity >95% based on HPLC analysis, see Supporting Information). 1H NMR (400 MHz, CDCl3), 8.63 (d, J = 8.8 Hz, 1 H), 8.59 (d, J = 8.4 Hz, 1 H), 8.54 (d, J = 8.8 Hz, 2 H), 8.11–8.05 (m, 3 H), 7.71–7.64 (m, 2 H), 7.50–7.38 (m, 3 H), 7.30 (d, J = 8.4 Hz, 1 H), 7.26 (d, J = 7.6 Hz, 2 H), 7.00 (d, J = 2.0 Hz, 1 H), 6.62 (dd, J = 2.0 Hz, 8.4 Hz, 1 H), 6.36 (d, J = 8.8 Hz, 1 H), 2.92 (s, 6 H), 2.91 (s, 6 H), 2.33 (s, 3H). 13C NMR (100 MHz, CDCl3), δ 151.9, 151.8, 147.7, 146.2, 140.0, 139.8, 132.0, 131.9, 131.8, 131.2, 131.1, 130.7, 130.1 (2 C), 129.8, 129.7, 129.0, 128.9, 122.9, 121.8, 120.6, 120.3, 119.6, 119.5, 117.7, 115.6 (2 C), 114.2, 113.9, 113.8, 104.8, 45.4, 11.2. HRMS (TOF-ES+): m/z [M + 1]+ calcd for C37H33N3O6S2, 680.1889; found, 680.1891.
Cell Culture and Reagents
All cell lines, except for SUM1315MO2 and SUM149PT, were obtained from the Tissue Culture Shared Resource of Georgetown University Medical Center. The reagents for cell culture were purchased from Invitrogen (Carlsbad, CA, USA), Lonza (Basel, Switzerland), or Cellgro (Manassas, VA, USA). T47D, HS578T, MDA-MB-436, MDA-MB-453, MDA-MB-468, BT-549, A431, and PC-3 cells were maintained in DMEM containing 10% (v/v) heat-inactivated FBS (HI-FBS; HyClone, Logan, UT, USA) and 100 units/mL penicillin/streptomycin. MDA-MB-231 and MCF7 cells were maintained in DMEM with 5% HI-FBS and 100 units/mL penicillin/streptomycin. HCC1937 cells were grown in RPMI with 10% HI-FBS, 100 units/mL penicillin/streptomycin, and 1% HEPES. A549 lung cell line (ATCC, Manassas, VA) was cultured in RPMI-1640 with l-glutamine (Mediatech Inc., Herdon, VA), 5% FBS, and 2.5 mM l-glutamine. SUM1315MO2 and SUM149PT were maintained according to manufacturer’s recommendations (Asterand, Detroit, MI, USA). Viable cells were monitored by the Luna automated cell counter (Logos Biosystems, Gyunggi-do, Korea). AZD1480 was purchased from Selleck Chemicals (Houston, TX, USA), and CP690550 was obtained from LC Laboratories (Woburn, MA, USA). Stock solutions of compounds were made at 10 mM concentration in dimethyl sulfoxide (DMSO) and stored at −20 °C in small aliquots. Mahanine was a kind gift from Dr. Samir Bhattacharya and Dr. Bikas C. Pal (Indian Institute of Chemical Biology, Kolkata, India) and dissolved in ethanol. IL-6 was purchased from Sigma-Aldrich (St. Louis, MO, USA).
Multiphoton Laser Imaging
After incubation of slides with a fibronectin/cortactin solution for 30 min, MDA-MB-231 cells were plated onto the slides and incubated overnight. The cells were then treated with a 10 μM solution of 12 or DMSO control and incubated for 1 h before washing with PBS. The slides were fixed with a 4% formaldehyde solution and washed with PBS. The compound was excited at 725 nm with a multiphoton laser and imaged with a 500–550 nm filter.
Plasmid DNAs and Reporter Gene Assay
Transfection and reporter gene assay was performed as described.(61) APRE-Luc was described previously(51) and kindly provided by Dr. T. Hirano (Osaka University, Osaka, Japan). STAT3C expression DNA was described previously(50) and kind gift from Dr. J. Bromberg (Memorial Sloan–Kettering Cancer Center, Weill Cornell Medical College, New York, NY).
Cell Proliferation Assay
MTT (3-(4,5)-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay was performed as described previously.(62) The half-maximal inhibitory concentration (EC50) and the concentration for 50% of maximal inhibition of cell proliferation (GI50) values were calculated by CompuSyn software V1.0 (ComboSyn, Paramus, NJ).
Western Blot Analysis
Western blot analysis was performed on cell and tissue lysates as described.(62) Antibodies used in this study were as follows: phospho-STAT3 (Y705) (no. 9145) and STAT3 (no. 4904) from Cell Signaling (Danvers, MA), Cyclin D1 (sc-717) from Santa Cruz Biotechnologies (Santa Cruz, CA), β-actin (A1978) from Sigma-Aldrich (St. Louis, MO, USA), and GAPDH (no. 3783) from ProsciΨ (Poway, California, USA).
Reverse Transcription-PCR (RT-PCR)
The RT-PCR was performed as described.(63) The primers used in this study were synthesized from Sigma Genosys (St. Louise, MO, USA) or Bioneer (Seoul, Korea) with the following sequences: survivin, forward 5′-CCA CTG AGA ACG AGC CAG AC-3′ and reverse 5′-GCA CTT TCT TCG CAG TTT CC-3′; cyclin D1, forward 5′-CCG TCC ATG CGG AAG ATC-3′ and reverse 5′-ATG GCC AGC GGG AAG AC-3′; PTPN6, forward 5′-TCT CTT GCA AGC ATT GGC AAG GTC-3′ and reverse 5′-TGC CAG GAT TCA AAC CCA GAC AGT-3′; GAPDH, forward 5′-GTA TGA CAA CGA ATT TGG CTA CAG-3′ and reverse 5′-AGC ACA GGG TAC TTT ATT GAT GGT-3′.
Transfection of siRNA
To knockdown the PTPN6 expression, transfection of siRNA was performed as described previously.(64) In brief, cells in 6-well plates were transfected with 100 nM/well of siRNA by Lipofectamine 2000 reagent (Invitrogen) in serum free media. Four hours after transfection, an equal volume of normal growth media containing 20% FBS was added to each well and the cells were further incubated for 3 days. Transfected cells were further treated with either DMSO or 12 for 24 h before harvest, and then Western blot analysis was performed. Densitometry analysis was performed by ImageJ.(65) The siRNAs were purchased from Bioneer (Seoul, Korea) with following sequences: control-siRNA, 5′-GAC GAG CGG CAC GUG CAC AUU-3′; PTPN6-2, 5′-CAG UUC AUU GAA ACC ACU A(dTdT)-3′; and PTPN6-4, 5′-GAG AAC GCU AAG ACC UAC A(dTdT)-3′.
Animals
Balb/c mice and athymic balb/c nude mice were purchased from the NCI. Animals were housed five per cage with microisolater tops and provided food (Purina mice chow) and water ad libitum. The light cycle was regulated automatically (12 h light/dark cycle), and temperature was maintained at 23 ± 1 °C. All animals were allowed to acclimate to this environment for 1 week prior to experimental manipulations. The Georgetown University Animal Care and Use Committee approved all animal studies (protocol no. 11-029) in accordance with the guideline adopted by the National Institutes of Health.
In Vivo Xenograft Study
The stock solution of compound 12 was dissolved in DMSO at 200 mg/mL. The working solution was prepared in a Cremophore and PEG400 mixture (Cremophore 10% and PEG400 3% and 1× PBS 87%) to give a 1 mg/mL solution. Male and female athymic balb/c nude mice (18–22 g) were injected with human A549 lung or SUM149 breast cancer cells (3 × 106 cells in a volume of 0.3 mL) into the subcutaneous tissue of the right auxiliary region of the mice. Three days after the tumor cell inoculation, the mice were randomly sorted into three groups with five mice per group. The tumor-bearing mice were either given an ip injection 10 mg/kg of compound 12 or the vehicle only once a day on every other day. The treatment was initiated when the tumor burden of mice reached around 50 mm3. The tumor size was measured by caliper three times a week to document tumor growth and calculated by the formula: length × width × width/2, and the body weight was measured and recorded. The therapeutic efficacy was evaluated based on body weight loss, tumor growth inhibition [determined by using calipers and calculated with formula: tumor inhibit rate (%) = (tumor volCon – tumor volTre)/tumor volCon].
Statistical Analysis
To compare two groups of interest, the two-tailed Student’s t test was applied for statistical analysis. * indicates P < 0.05, ** indicates P < 0.01, and *** indicates P < 0.001.
Supporting Information
1H and 13 C NMR spectra of new compounds and HPLC purity analysis of compounds 12 and 13. This material is available free of charge via the Internet at http://pubs.acs.org.
⊥Author Contributions
S.H., Y.W.Y., and H.J.K. contributed equally to this work.
The authors declare the following competing financial interest(s): Patent applications have been filed by Georgetown University on the behalf of the inventors that are listed as authors in this work.
Acknowledgment
We thank Tissue Culture Shared Resources supported by P30-CA051008-18 (LCCC Cancer Center Support Grant) at Georgetown University Medical Center. We are grateful to Dr. T. Hirano and Dr. J. Bromberg for kind gifts of plasmid DNAs. We also thank Dr. Samir Bhattacharya and Dr. Bikas C. Pal for providing purified mahanine. This work was supported by the Center for Drug Discovery at Georgetown University Medical Center, Susan G. Komen for the Cure (FAS0703858), and the Lombardi Comprehensive Cancer Center, Georgetown University (P30-CA051008).
| Abbreviations Used | |
| Dansyl | 5-(dimethylamino)naphthalene |
| TLC | thin layer chromatography |
| DCB | dichloronenzene |
| DCM | dichloromethane |
| TEA | triethylamine |
| NMR | nuclear magnetic resonance |
| MTT | 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide |
| PEG | polyethylene glycol |
| GI50 | concentration of drug needed to inhibit cell growth by 50% |
| EC50 | half-maximal effective concentration of drug |
| LD50 | dose where 50% of test subjects exposed would be killed |
| CT | Control |
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, J.; Darnell, J. E., Jr. The role of STATs in transcriptional control and their impact on cellular function Oncogene 2000, 19, 2468– 2473[Crossref], [PubMed], [CAS]8.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXktVGhsb4%253D&md5=b504533a9132be2af9c0a0a827b93063Bromberg, Jacqueline; Darnell, James E, Jr.Oncogene (2000),A review with 68 refs. The STAT proteins (Signal Transducers and Activators of Transcription), were identified in the last decade as transcription factors which were crit. in mediating virtually all cytokine driven signaling. These proteins are latent in the cytoplasm and become activated through tyrosine phosphorylation which typically occurs through cytokine receptor assocd. kinases (JAKs) or growth factor receptor tyrosine kinases. Recently a no. of non-receptor tyrosine kinases (for example src and abl) have been found to cause STAT phosphorylation. Phosphorylated STATs form homo- or hetero-dimers, enter the nucleus and working coordinately with other transcriptional co-activators or transcription factors lead to increased transcriptional initiation. In normal cells and in animals, ligand dependent activation of the STATs is a transient process, lasting for several minutes to several hours. In contrast, in many cancerous cell lines and tumors, where growth factor dysregulation is frequently at the heart of cellular transformation, the STAT proteins (in particular Stats 1, 3 and 5) are persistently tyrosine phosphorylated or activated. The importance of STAT activation to growth control in expts. using anti-sense mols. or dominant neg. STAT protein encoding constructs performed in cell lines or studies in animals lacking specific STATs strongly indicate that STATs play an important role in controlling cell cycle progression and apoptosis. Stat1 plays an important role in growth arrest, in promoting apoptosis and is implicated as a tumor suppressor; while Stats 3 and 5 are involved in promoting cell cycle progression and cellular transformation and preventing apoptosis. Many questions remain including: (1) a better understanding of how the STAT proteins through assocn. with other factors increase transcription initiation; (2) a more complete definition of the sets of genes which are activated by different STATs and (3) how these sets of activated genes differ as a function of cell type. Finally, in the context of many cancers, where STATs are frequently persistently activated, an understanding of the mechanisms leading to their constitutive activation and defining the potential importance of persistent STAT activation in human tumorigenesis remains. - 9.Mankan
, A. K.; Greten, F. R. Inhibiting signal transducer and activator of transcription 3: rationality and rationale design of inhibitors Expert Opin. Invest. Drugs 2011, 20, 1263– 1275[Crossref], [PubMed], [CAS]9.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhtVWntbbP&md5=648ce046310a6fb7a3203e9be9367376Mankan, Arun K.; Greten, Florian R.Expert Opinion on Investigational Drugs (2011),A review. Signal transducer and activator of transcription 3 (STAT3) controls a key signaling pathway in the development of many malignant diseases. Several genetic studies have proven its central role in the regulation of apoptosis, proliferation, angiogenesis and immune responses making it an attractive target for cancer therapy.Areas covered: This article addresses the role of STAT3 in immune response modulation and highlights the contribution of STAT3 in inflammation-mediated tumorigenesis. We also review the rationale to use novel STAT3 inhibitors and list some of these inhibitors such as STA-21, IS3 295, S3I- M2001 and small mol. JAK2 inhibitors AZD1480 and AZ960 that were found to be efficient against tumors. We summarize the efforts that were made so far in identifying promising compds. and mention the barriers that need to be overcome for successful application of STAT3 inhibitors in clinics.Expert opinion: STAT3 is an important target in tumor biol. based on its frequent activation in various tumors and its pleiotropic effects on different cell types. Screening large libraries of logically synthesized small mol. inhibitors is one way to rapidly generate many potential mols., which can then be tested in different biol. relevant models. The stage is, therefore, set for the identification and development of novel STAT3 inhibitors that will, in the very near future, enter the clin. realm. - 10.Horiguchi
, A.; Oya, M.; Shimada, T.; Uchida, A.; Marumo, K.; Murai, M. Activation of singal transducer and activator 3 in renal cell carcinoma: a study of incidence and its association with pathological features and clinical outcome J. Urol. 2002, 168, 762– 765[Crossref], [PubMed], [CAS]10.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38XmtFWhurw%253D&md5=a8e5fe3f02f14b11202b78d40a42945bHoriguchi, Akio; Oya, Mototsugu; Shimada, Tetsuya; Uchida, Atsushi; Marumo, Ken; Murai, MasaruJournal of Urology (Hagerstown, MD, United States) (2002),Purpose: Signal transducer and activator of transcription 3 (STAT3) is known to have an important role in cytokine and growth factor signaling pathways. In various types of human malignant tumors STAT3 has been shown to be constitutively activated due to aberrant prodn. of cytokines and growth factors. The authors examd. the presence of STAT3 activation and its assocn. with pathol. features and clin. outcome in renal cell carcinoma cases. Materials and Methods: the authors examd. 48 paraffin embedded renal cell carcinoma specimens and corresponding nonneoplastic kidney tissues for the activation status of STAT3 on immunohistochem. using anti-phospho-specific (p)-STAT3 antibody, which recognizes only activated STAT3. Based on the percent of cells with pos. nuclear staining the activation status of STAT3 was detd. and categorized into 2 groups, including low - less than 10% and high - 90% or more tumor cells pos. The assocns. of the activation status of STAT3 with pathol. features and clin. outcome were analyzed. Results: Of 48 tumors 24 (50%) demonstrated high levels of nuclear immunostaining for p-STAT3, while the other 24 (50%) showed low levels. Adjacent nonneoplastic kidney tissues showed only little immunostaining for p-STAT3. A significant assocn. of high levels of p-STAT3 with metastasis was obsd. No significant assocns. of p-STAT3 immunostaining with pathol. stage or grade were obsd. A high level of p-STAT3 was a significant indicator of a poor prognosis on univariate and multivariate anal. Conclusions: the authors' results indicate a high frequency of STAT3 activation in renal cell carcinoma, esp. in metastatic disease. STAT3 activation was an independent prognostic variable in renal cell carcinoma cases. The authors' results strongly suggest that the activation of STAT3 contributes to the development and progression of renal cell carcinoma. - 11.Takemoto
, S.; Ushijima, K.; Kawano, K.; Yamaguchi, T.; Terada, a; Fujiyoshi, N.; Nishio, S.; Tsuda, N.; Ijichi, M.; Kakuma, T.; Kage, M.; Hori, D.; Kamura, T. Expression of activated signal transducer and activator of transcription-3 predicts poor prognosis in cervical squamous-cell carcinoma Br. J. Cancer 2009, 101, 967– 972[Crossref], [PubMed], [CAS]11.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXhtV2qsrnN&md5=1593e05bfde3988ed119127a3860653dTakemoto, S.; Ushijima, K.; Kawano, K.; Yamaguchi, T.; Terada, A.; Fujiyoshi, N.; Nishio, S.; Tsuda, N.; Ijichi, M.; Kakuma, T.; Kage, M.; Hori, D.; Kamura, T.British Journal of Cancer (2009),Background: Stat3 is a member of the Janus-activated kinase/STAT signalling pathway. It normally resides in the cytoplasm and can be activated through phosphorylation. Activated Stat3 (p-Stat3) translocates to the nucleus to activate the transcription of several mols. involved in cell survival and proliferation. The constitutive activation of Stat3 has been shown in various types of malignancies, and its expression has been reported to indicate a poor prognosis. However, the correlation between the constitutive activation of Stat3 and the prognosis of cervical cancer patients has not been reported. Methods: The immunohistochem. anal. of p-Stat3 expression was performed on tissues from 125 cervical squamous-cell carcinoma patients who underwent extended hysterectomy and pelvic lymphadenectomy, and the assocn. of p-Stat3 expression with several clinicopathol. factors and survival was investigated. Results: Pos. p-Stat3 expression was obsd. in 71 of 125 (56.8%) cases and was significantly correlated with lymph node metastasis, lymph vascular space invasion, and large tumor diam. (>4 cm) by Fisher's exact test. Kaplan-Meier survival anal. showed that p-Stat3 expression was statistically indicative of a poor prognosis for overall survival (P=0.006) and disease-free survival (P=0.010) by log-rank test. Conclusion: These data showed that p-Stat3 expression in cervical cancer acts as a predictor of poor prognosis. - 12.Zhang
, X. L.; Yue, P. B.; Page, B. D. G.; Li, T. S.; Zhao, W.; Namanja, A. T.; Paladino, D.; Zhao, J. H.; Chen, Y.; Gunning, P. T.; Turkson, J. Orally bioavailable small-molecule inhibitor of transcription factor stat3 regress humane breast and lung cancer xenografts Proc. Natl. Acad. Sci. U. S. A. 2012, 109, 9623– 9628[Crossref], [PubMed], [CAS]12.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XptlWjtLo%253D&md5=4da78174a7c00eacc1f1a1ce553c5aa7Zhang, Xiaolei; Yue, Peibin; Page, Brent D. G.; Li, Tianshu; Zhao, Wei; Namanja, Andrew T.; Paladino, David; Zhao, Jihe; Chen, Yuan; Gunning, Patrick T.; Turkson, JamesProceedings of the National Academy of Sciences of the United States of America (2012),Computer-aided lead optimization derives a unique, orally bioavailable inhibitor of the signal transducer and activator of transcription (Stat)3 Src homol. 2 domain. BP-1-102 binds Stat3 with an affinity (KD) of 504 nM, blocks Stat3-phospho-tyrosine (pTyr) peptide interactions and Stat3 activation at 4-6.8 μM, and selectively inhibits growth, survival, migration, and invasion of Stat3-dependent tumor cells. BP-1-102-mediated inhibition of aberrantly active Stat3 in tumor cells suppresses the expression of c-Myc, Cyclin D1, Bcl-xL, Survivin, VEGF, and Kruppel-like factor 8, which is identified as a Stat3 target gene that promotes Stat3-mediated breast tumor cell migration and invasion. Treatment of breast cancer cells with BP-1-102 further blocks Stat3-NF-κB cross-talk, the release of granulocyte colony-stimulating factor, sol. intercellular adhesion mol. 1, macrophage migration-inhibitory factor/glycosylation-inhibiting factor, interleukin 1 receptor antagonist, and serine protease inhibitor protein 1, and the phosphorylation of focal adhesion kinase and paxillin, while enhancing E-cadherin expression. I.v. or oral gavage delivery of BP-1-102 furnishes micromolar or microgram levels in tumor tissues and inhibits growth of human breast and lung tumor xenografts. - 13.Lin
, L.; Amin, R.; Gallicano, G. I.; Glasgow, E.; Jogunoori, W.; Jessup, J. M.; Zasloff, M.; Marshall, J. L.; Shetty, K.; Johonson, L.; Mishra, L.; He, A. R. The STAT3 inhibitor NSC 78459 is effective in hepatocellular cancers with disrupted TGF-β signaling Oncogene 2009, 28, 961– 972[Crossref], [PubMed], [CAS]13.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXitFKltLc%253D&md5=5807decfd3769d71b1f3ad93516d7647Lin, L.; Amin, R.; Gallicano, G. I.; Glasgow, E.; Jogunoori, W.; Jessup, J. M.; Zasloff, M.; Marshall, J. L.; Shetty, K.; Johnson, L.; Mishra, L.; He, A. R.Oncogene (2009),Hepatocellular carcinoma (HCC) is the third leading cause of cancer deaths worldwide, with few effective therapeutic options for advanced disease. At least 40% of HCCs are clonal, potentially arising from STAT3+, NANOG+ and OCT3/4+ liver progenitor/stem cell transformation, along with inactivation of transforming growth factor-beta (TGF-β) signaling. Here the authors report significantly greater signal transducer and activator of transcription 3 (STAT3) and tyrosine phosphorylated STAT3 in human HCC tissues (P<0.0030 and P<0.0455, resp.) than in human normal liver. Further, in HCC cells with loss of response to TGF-β, NSC 74859, a STAT3-specific inhibitor, markedly suppresses growth. In contrast, CD133+ status did not affect the response to STAT3 inhibition: both CD133+ Huh-7 cells and CD133- Huh-7 cells are equally sensitive to NSC 74859 treatment and STAT3 inhibition, with an IC50 of 100 μM. Thus, the TGF-β/beta2 spectrin (β2SP) pathway may reflect a more functional 'stem/progenitor' state than CD133. Furthermore, NSC 74859 treatment of Huh-7 xenografts in nude mice significantly retarded tumor growth, with an ED of only 5 mg/kg. Moreover, NSC 74859 inhibited tyrosine phosphorylation of STAT3 in HCC cells in vivo. The authors conclude that inhibiting interleukin 6 (IL6)/STAT3 in HCCs with inactivation of the TGF-β/β2SP pathway is an effective approach in management of HCCs. Thus, IL6/STAT3, a major signaling pathway in HCC stem cell renewal and proliferation, can provide a novel approach to the treatment of specific HCCs. - 14.Sen
, M.; Tosca, P. J.; Zwayer, C.; Ryan, M. J.; Johnson, J. D.; Knostman, K. A. Lack of toxicity of a STAT3 decoy oligonucleotide Cancer Chemother. Pharmacol. 2009, 63, 983– 995[Crossref], [PubMed], [CAS]14.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXjslOlurw%253D&md5=ac604401fe84142a1568bf2d58171c22Sen, Malabika; Tosca, Patricia J.; Zwayer, Christa; Ryan, Michael J.; Johnson, Jerry D.; Knostman, Katherine A. B.; Giclas, Patricia C.; Peggins, James O.; Tomaszewski, Joseph E.; McMurray, Timothy P.; Li, Changyou; Leibowitz, Michael S.; Ferris, Robert L.; Gooding, William E.; Thomas, Sufi M.; Johnson, Daniel E.; Grandis, Jennifer R.Cancer Chemotherapy and Pharmacology (2009),STAT3 overexpression has been detected in several cancers including head and neck squamous cell carcinoma (HNSCC). Previous studies using intratumoral administration of a STAT3 decoy oligonucleotide that abrogates STAT3-mediated gene transcription in preclin. cancer models have demonstrated antitumor efficacy. This study was conducted to observe the toxicity and biol. effects of the STAT3 decoy in a nonhuman primate model, in anticipation of initiating a clin. trial in HNSCC patients. Three study groups (2 monkeys/sex/group) were administered a single i.m. injection of low dose of STAT3 decoy (0.8 mg total dose/monkey), high dose of STAT3 decoy (3.2 mg total dose/monkey), or vehicle control (PBS alone) on day 1 and necropsies were performed on days 2 and 15 (one monkey/sex/group/day). Low and high doses of the decoy were administered in the muscle in a vol. of 0.9 mL. Tissue and blood were harvested for toxicol. and biol. analyses. Upon observation, the STAT3 decoy-treated animals exhibited behavior that was similar to the vehicle control group. Individual animal body wts. remained within 1% of pretreatment wts. throughout the study. Hematol. parameters were not significantly different between the control and the treatment groups. Clin. chem. fluctuations were considered within normal limits and were not attributed to the STAT3 decoy. Assessment of complement activation breakdown product (Bb) levels demonstrated no activation of the alternative pathway of complement in any animal at any dose level. At necropsy, there were no gross or microscopic findings attributed to STAT3 decoy in any organ examd. STAT3 target gene expression at the injection site revealed decreased Bcl-XL and cyclin D1 expression levels in the animals treated with high dose of STAT3 decoy compared to the animals injected with low dose of STAT3 decoy or the vehicle as control. Based on these findings, the no-observable-adverse-effect level (NOAEL) was >3.2 mg/kg when administered as a single dose to male and female Cynomolgus monkeys. Plans are underway to test the safety and biol. effects of intratumoral administration of the STAT3 decoy in HNSCC patients. - 15.Turkson
, J.; Jove, R. STAT protein: a novel molecular target for cancer drug discovery Oncogene 2000, 19, 6613– 6626[Crossref], [PubMed], [CAS]15.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3MXhsF2rsro%253D&md5=2011fb621ef70547ddbbef2ed253df98Turkson, James; Jove, RichardOncogene (2000),A review with 101 refs. Signal Transducers and Activators of Transcription (STATs) are a family of cytoplasmic proteins with roles as signal messengers and transcription factors that participate in normal cellular responses to cytokines and growth factors. Frequently, however, abnormal activity of certain STAT family members, particularly Stat3 and Stat5, is assocd. with a wide variety of human malignancies, including hematol., breast, head and neck, and prostate cancers. Application of mol. biol. and pharmacol. tools in disease-relevant models has confirmed Stat3 as having a causal role in oncogenesis, and provided validation of Stat3 as a target for cancer drug discovery and therapeutic intervention. Furthermore, a constitutively-active mutant form of Stat3 is sufficient to induce oncogenic transformation of cells, which form tumors in vivo. Constitutive activation of Stat3 signaling is accompanied by upregulation of cyclin D1, c-Myc, and Bcl-x, changes consistent with subversion of normal cellular growth and survival control mechanisms. Block of constitutive Stat3 signaling results in growth inhibition and apoptosis of Stat3-pos. tumor cells in vitro and in vivo. The obsd. dependence of certain tumors on constitutive Stat3 signaling for growth and survival has wide implications for cancer therapy, offering the potential for preferential tumor cell killing. This review evaluates constitutive Stat3 activation as a "cancer-causing" factor, and proposes a no. of mol. strategies for targeting Stat3 signaling for therapeutic intervention. - 16.Debnath
, B.; Xu, S.; Neamati, N. Small molecule inhibitors of signal transducer and activator of transcription 3 (Stat3) protein J. Med. Chem. 2012, 55, 6645– 6668[ACS Full Text
], [CAS]16.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38Xnsl2msrc%253D&md5=f5f3ac9f8fc2326c75edd2a9c136275bDebnath, Bikash; Xu, Shili; Neamati, NouriJournal of Medicinal Chemistry (2012),A review. Signal transducer and activator of transcription 3 (Stat3) is a transcription factor belonging to the STAT protein family, members of which regulate a wide variety of biol. processes, including cell growth, proliferation, survival, differentiation, development, movement and inflammation in response to extracellular signaling mols. such as cytokines and growth factors. Receptor assocd. tyrosine kinases phosphorylate Stat3 protein at Tyr705 and enable it to dimerize and subsequently enter the nucleus where it activates the transcription of its target genes. Accumulating evidence has established Stat3's role in oncogenesis, cardiovascular diseases, rheumatoid arthritis, fibrotic, and Alzheimer's diseases. Thus, Stat3 inhibition is a promising therapeutic approach not only for cancer, but also for other diseases. Although Stat3 has been considered as an important therapeutic target, progress in discovery and development of effective and specific Stat3 inhibitors remains slow. In this article we summarize Stat3 inhibitors published between 2006 and 2011. New strategies along with novel technologies are required to achieve breakthrough in the development of Stat3 inhibitors that can be translated into clin. use. - 17.Germain
, D.; Frank, D. A. Targeting the cytoplasmic and nuclear functions of signal transducer and activitors of transcription 3 for cancer therapy Clin. Cancer Res. 2007, 13, 5665– 5669[Crossref], [PubMed], [CAS]17.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXhtFSntLzK&md5=7c06b8b398fa29f41b1e2049abbb04eaGermain, Doris; Frank, David A.Clinical Cancer Research (2007),A review. Signal transducers and activators of transcription (STAT) are a highly conserved family of transcription factors that are activated by phosphorylation in the cytoplasm, after which they translocate to the nucleus to regulate gene expression. Among the seven STATs, STAT3 is of particular interest due to its constitutive phosphorylation in a large proportion of human cancers and its ability to induce neoplastic transformation. Inhibition of STAT3 can reverse tumor growth in exptl. systems while having few effects in normal cells. These findings have implicated STAT3 as a potentially important target for therapeutic intervention. In addn. to its well-described role as a transcription factor, STAT3 has been found recently to have important effects in the cytoplasm. Collectively, these functions of STAT3 directly contribute to tumorigenesis, invasion, and metastasis. Given the potential importance of STAT3 as a target for cancer therapy, mols. have been developed that can block STAT3 function at a variety of steps. These drugs show promise as anticancer agents in model systems of a variety of common human cancers. Thus, elucidating the functions of STAT3 and developing agents to inhibit this protein remain important scientific and clin. challenges. - 19.Santos
, F. P. S.; Kantarjian, H. M.; Jain, N.; Manshouri, T.; Thomas, D. A.; Garcia-Manero, G.; Kennedy, D.; Estrov, Z.; Cortes, J.; Verstovsek, S. Phase 2 study of CEP-701, an orally available JAK2 inhibitor, in patients with primary or post-polycythemia vera/essential thrombocythemia myelofibrosis Blood 2010, 115, 1131– 1136[Crossref], [PubMed], [CAS]19.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXisFyrt7k%253D&md5=02b4a0fa1af657f0552cd7a0f11250caSantos, Fabio P. S.; Kantarjian, Hagop M.; Jain, Nitin; Manshouri, Taghi; Thomas, Deborah A.; Garcia-Manero, Guillermo; Kennedy, Debra; Estrov, Zeev; Cortes, Jorge; Verstovsek, SrdanBlood (2010),Few treatment options exist for patients with myelofibrosis (MF), and their survival is significantly shortened. Activating mutation of the JAK2 tyrosine kinase (JAK2V617F) is found in approx. 50% of MF patients. CEP-701 is a tyrosine kinase inhibitor that inhibits JAK2 in in vitro and in vivo expts. We conducted a phase 2 clin. study of CEP-701 in 22 JAK2V617F-pos. MF patients (80 mg orally twice daily), and 6 (27%) responded by International Working Group criteria (clin. improvement in all cases): redn. in spleen size only (n = 3), transfusion independency (n = 2), and redn. in spleen size with improvement in cytopenias (n = 1). Median time to response was 3 mo, and duration of response was more than or equal to 14 mo. No improvement was seen in bone marrow fibrosis or JAK2V617F allele burden. Phosphorylated STAT3 levels decreased from baseline in responders while on therapy. Eight patients (36%) experienced grade 3 or 4 toxicity, and 6 (27%) required dose redn. Main side effects were myelosuppression (grade 3 or 4 anemia, 14%; and thrombocytopenia, 23%) and gastrointestinal disturbances (diarrhea, any grade, 72%; grade 3 or 4, 9%; nausea, grade 1 or 2 only, 50%; vomiting, grade 1 or 2 only, 27%). In conclusion, CEP-701 resulted in modest efficacy and mild but frequent gastrointestinal toxicity in MF patients. - 20.Chen
, H.; Yang, Z.; Ding, C.; Chu, L.; Zhang, Y.; Terry, K.; Liu, H.; Shen, Q.; Zhou, J. Discovery of O-alkylamino tethered niclosamide derivatives as potent and orally bioavailable anticancer agents ACS Med. Chem. Lett. 2013, 4, 180– 185[ACS Full Text], [CAS]20.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXns1ejug%253D%253D&md5=a481986f86800fc39f24a6af12c46a8eChen, Haijun; Yang, Zhengduo; Ding, Chunyong; Chu, Lili; Zhang, Yusong; Terry, Kristin; Liu, Huiling; Shen, Qiang; Zhou, JiaACS Medicinal Chemistry Letters (2013),Niclosamide has been identified to potently inhibit the activation, nuclear translocation, and transactivation of STAT3. Nevertheless, the poor aq. soly. and bioavailability of niclosamide have hindered its further clin. development for cancer therapy. To discover new mols. with enhanced druglike properties, a series of novel O-alkylamino-tethered derivs. of niclosamide have been designed, synthesized, and biol. evaluated. Among them, compd. I·HCl (HJC0152) has been demonstrated to significantly suppress MDA-MB-231 xenograft tumor growth in vivo (i.p. and po), indicating its great potential as efficacious and orally bioavailable therapeutics for human cancer. - 21.Chen
, H.; Yang, Z.; Ding, C.; Chu, L.; Zhang, Y.; Terry, K.; Liu, H.; Shen, Q.; Zhou, J. Fragment-based drug design and identification of HJC0123, a novel orally bioavailable STAT3 inhibitor for cancer therapy Eur. J. Med. Chem. 2013, 62, 498– 507[Crossref], [PubMed], [CAS]21.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXlslWmsb4%253D&md5=218e2dad165f5188faa353bbc6c9ab67Chen, Haijun; Yang, Zhengduo; Ding, Chunyong; Chu, Lili; Zhang, Yusong; Terry, Kristin; Liu, Huiling; Shen, Qiang; Zhou, JiaEuropean Journal of Medicinal Chemistry (2013),Fragment-based drug design is a promising approach for the generation of lead mols. with enhanced activity and esp. drug-like properties against therapeutic targets. Herein, the authors report the fragment-based drug design, systematic chem. synthesis and pharmacol. evaluation of novel scaffolds as potent anticancer agents by utilizing six privileged fragments from known STAT3 inhibitors. Several new mols. such as compds. HJC0123, 5-Chloro-N-(1,1-dioxo-1H-1λ6-benzo[b]thiophen-6-yl)-2hydroxybenzamide, N-(1,1-Dioxo-1H-1λ66-benzo[b]thiophen-6-yl)-2,6-diphenylisonicotinamide that may act as advanced chem. leads have been identified. The most potent compd. HJC0123 has demonstrated to inhibit STAT3 promoter activity, downregulate phosphorylation of STAT3, increase the expression of cleaved caspase-3, inhibit cell cycle progression and promote apoptosis in breast and pancreatic cancer cells with low micromolar to nanomolar IC50 values. Furthermore, compd. HJC0123 significantly suppressed estrogen receptor (ER)-neg. breast cancer MDA-MB-231 xenograft tumor growth in vivo, indicating its great potential as an efficacious and orally bioavailable drug candidate for human cancer therapy. - 22.Miyoshi
, K.; Takaishi, M.; Nakajima, K.; Ikeda, M.; Kanda, T.; Tarutani, M.; Iiyama, T.; Asao, N.; DiGiovanni, J.; Sano, S. Stat3 as a therapeutic target for the treatment of psoriasis: a clinical feasibility study with STA-21, a Stat3 Inhibitor J. Invest. Dermatol. 2011, 131, 108– 117[Crossref], [PubMed], [CAS]22.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXhsFGrsb3F&md5=5bc8fa76b394459a7f06e6558c75eeddMiyoshi, Ken; Takaishi, Mikiro; Nakajima, Kimiko; Ikeda, Mitsunori; Kanda, Takashi; Tarutani, Masahito; Iiyama, Tatsuo; Asao, Naoki; Di Giovanni, John; Sano, ShigetoshiJournal of Investigative Dermatology (2011),Epidermal keratinocytes in psoriatic lesions are characterized by activated Stat3, and increased levels of cytokines and growth factors that promote Stat3 activation have been found within psoriatic lesions. K5.Stat3C transgenic mice, in which keratinocytes express a constitutively active Stat3, develop psoriasis-like skin lesions. In this study, we examd. whether STA-21, a small Stat3 inhibitor, could be useful in ameliorating the skin lesions not only in the model mouse but also in human psoriasis. Treatment with STA-21 markedly inhibited the cytokine-dependent nuclear translocation of Stat3 in normal human keratinocytes in vitro. Keratinocyte proliferation was inhibited by STA-21 in a dose-dependent manner through downregulation of c-Myc and cyclin D1, whereas involucrin, transglutaminase 1, and keratin 10 levels were upregulated. Topical application of STA-21 abolished the generation of skin lesions in K5.Stat3C mice. Finally, we treated psoriasis patients with STA-21-contg. ointment in a nonrandomized study. Psoriatic lesions in six of the eight patients showed improvement after topical STA-21 treatment for 2 wk. Therefore, we conclude that targeting Stat3 may lead to a therapy for psoriasis. Journal of Investigative Dermatol. (2011) 131, 108-117; doi:10.1038/jid.2010.255; published online 2 Sept. 2010. - 23.Newman
, D. J.; Cragg, G. M. Natural products as sources of new drug discover over the last 25 years J. Nat. Prod. 2007, 70, 461– 477[ACS Full Text], [CAS]23.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXhvFKrsro%253D&md5=9fc0909750367ccd3b97589aaf087cdfNewman, David J.; Cragg, Gordon M.Journal of Natural Products (2007),A review. This review is an updated and expanded version of two prior reviews that were published in this journal in 1997 and 2003. In the case of all approved agents the time frame has been extended to include the 251/2 years from 01/1981 to 06/2006 for all diseases worldwide and from 1950 (earliest so far identified) to 06/2006 for all approved antitumor drugs worldwide. We have continued to utilize our secondary subdivision of a "natural product mimic" or "NM" to join the original primary divisions. From the data presented, the utility of natural products as sources of novel structures, but not necessarily the final drug entity, is still alive and well. Thus, in the area of cancer, over the time frame from around the 1940s to date, of the 155 small mols., 73% are other than "S" (synthetic), with 47% actually being either natural products or directly derived therefrom. In other areas, the influence of natural product structures is quite marked, with, as expected from prior information, the antiinfective area being dependent on natural products and their structures. Although combinatorial chem. techniques have succeeded as methods of optimizing structures and have, in fact, been used in the optimization of many recently approved agents, we are able to identify only one de novo combinatorial compd. approved as a drug in this 25 plus year time frame. We wish to draw the attention of readers to the rapidly evolving recognition that a significant no. of natural product drugs/leads are actually produced by microbes and/or microbial interactions with the "host from whence it was isolated", and therefore we consider that this area of natural product research should be expanded significantly. - 25.Long
, B. H.; Rose, W. C.; Vyas, D. M.; Matson, J. A.; Forenza, S. Discovery of antitumor indolocarbazoles: rebeccamycin, NSC655649, and fluoroindolocarbazoles Curr. Med. Chem.: Anti-Cancer Agents 2002, 2, 255– 266[Crossref], [PubMed], [CAS]25.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38Xis1Omu78%253D&md5=288246997498715c1a5c0f7c0e7b172eLong, Byron H.; Rose, William C.; Vyas, Dolatrai M.; Matson, James A.; Forenza, SalvatoreCurrent Medicinal Chemistry: Anti-Cancer Agents (2002),A review. A fermn. directed product search for potential anticancer drugs conducted by Bristol-Myers in the 1970s and early 1980s resulted in the identification of a novel indolocarbazole (IC) rebeccamycin (RBM) as a potential drug development candidate. Subsequently, an analog program designed to impart distal site in vivo antitumor activity resulted in the discovery of diethylaminoethyl analog of RBM (DEAE-RBM), which is presently undergoing clin. evaluation as NSC 655649 and BMY-27557. Strong DNA intercalation is the primary mechanism of action of DEAE-RBM resulting in the potent catalytic inhibition of both topoisomerases I and II. Precursor feeding fermn. expts. with fluorine-substituted tryptophans yielded novel fluoroindolocarbazoles (FICs). These FICs were identified as targeting topoisomerase (topo) I in a mechanism-based screen and their action on topo I was confirmed by prodn. of topo I-mediated single-strand breaks in DNA at sites essentially identical to those induced by camptothecin. Topo I dependent cytotoxicity was demonstrated for specific FICs using a P388 and camptothecin-resistant P388/CPT45 pair of cell lines, the latter expresses little or no functional topo I. For example, topo I selectivity was greatest with 3,9-difluoro substituted FIC and was least significant and least cytotoxic with 4,8-difluoro substituted FIC. The review focuses on the discovery of the rebeccamycin class of compds. and their structure-activity relationships leading to the development of the clin. candidate BMY-27557 (NSC 655649), as well as the lead identification of the fluoroindolocarbazole class of compds. - 26.Prudhomme
, M. Rebeccamycin analogues as anti-cancer agents Eur. J. Med. Chem. 2003, 38, 123– 140[Crossref], [PubMed], [CAS]26.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3sXhs1eiur0%253D&md5=616458d7a26407e4012998f06ecc5e7bPrudhomme, MichelleEuropean Journal of Medicinal Chemistry (2003),This review reports the various total syntheses of rebeccamycin and structure-activity relationship studies on rebeccamycin analogs. Rebeccamycin, a microbial metabolite possessing a maleimide indolo[2,3-a]carbazole framework with a carbohydrate moiety attached to one of the indole nitrogens, is a well-known topoisomerase I inhibitor. Rebeccamycin analogs were prepd. either by semi-synthesis from the natural metabolite or by total synthesis. Different families of rebeccamycin analogs were obtained by modifications at the imide heterocycle, dechlorination and substitutions on the indole moieties, modifications of the sugar residue, construction of dimers, coupling the sugar unit to the second indole nitrogen, changing indolo[2,3-a]carbazole skeleton to indolo[2,3-c]carbazole, replacing one or both indole moieties by 7-azaindole units. The biol. activities of the rebeccamycin analogs are described. According to their chem. structure, the analogs can inhibit topoisomerase I and/or kinases. From the structure-activity relationships, some important rules were established. Several compds. exhibit stronger antiproliferative activities than the natural metabolite with IC50 values in the nanomolar range. Some analogs, esp. those possessing azaindole moieties, are much more selective than rebeccamycin toward the tumor cell lines tested. - 27.Arbiser
, J. L.; Govindarajan, B.; Battle, T. E.; Lynch, R.; Frank, D. A.; Ushio-Fukai, M.; Perry, B. N.; Stern, D. F.; Bowden, G. T.; Liu, A.; Klein, E.; Kolodziejski, P. J.; Eissa, N. T.; Hossain, C. F.; Nagle, D. G. Carbazole is a naturally occurring inhibitor of angiogenesis and inflammation isolated from antipsoriatic coal tar J. Invest. Dermatol. 2006, 126, 1396– 1402[Crossref], [PubMed], [CAS]27.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28Xls1Ght7s%253D&md5=958dcf099cde24e976b0496d355f618bArbiser, Jack L.; Govindarajan, Baskaran; Battle, Traci E.; Lynch, Rebecca; Frank, David A.; Ushio-Fukai, Masuko; Perry, Betsy N.; Stern, David F.; Bowden, G. Tim; Liu, Anquan; Klein, Eva; Kolodziejski, Pawel J.; Eissa, N. Tony; Hossain, Chowdhury F.; Nagle, Dale G.Journal of Investigative Dermatology (2006),Coal tar is one of the oldest and an effective treatment for psoriasis. Coal tar was directly applied to the skin, or used in combination with UV light as part of the Goeckerman treatment. The use of coal tar has caused long-term remissions in psoriasis, but has fallen out of favor because the treatment requires hospitalization and coal tar is poorly acceptable aesthetically to patients. Thus, detg. the active antipsoriatic component of coal tar is of considerable therapeutic interest. The authors fractionated coal tar into its components, and tested them using the SVR angiogenesis inhibitor assay. Treatment of SVR endothelial cells with coal tar fractions resulted in the isolation of a single fraction with antiangiogenic activity. The active antiangiogenic compd. in coal tar is carbazole. In addn. to antiangiogenic activity, carbazole inhibited the prodn. of inflammatory IL-15 by human mononuclear cells. IL-15 is elevated in psoriasis and is thought to contribute to psoriatic inflammation. Carbazole treatment also reduced activity of inducible nitric oxide synthase (iNOS), which is proinflammatory and elevated in psoriasis. The effect of carbazole on upstream pathways in human psoriasis was detd., and carbazole was shown to inhibit signal transducer and activator of transcription (stat)3-mediated transcription, which has been shown to be relevant in human psoriasis. IL-15, iNOS, and stat3 activation require the activation of the small GTPase rac for optimal activity. Carbazole was found to inhibit rac activation as a mechanism for its inhibition of downstream inflammatory and angiogenic pathways. Given its antiangiogenic and anti-inflammatory activities, carbazole is likely a major component of the antipsoriatic activity of coal tar. Carbazole and derivs. may be useful in the therapy of human psoriasis. - 28.Saturnino
, C.; Palladino, C.; Napoli, M.; Sinicropi, M. S.; Botta, A.; Sala, M.; Carcereri de Prati, A.; Novellino, E.; Suzuki, H. Synthesis and biological evaluation of new N-alkylcarbazole derivatives as STAT3 inhibitors: preliminary study Eur. J. Med. Chem. 2013, 60, 112– 119[Crossref], [PubMed], [CAS]28.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXjtVeju7w%253D&md5=87d8d33b28930068e31b4676f64df18aSaturnino, Carmela; Palladino, Chiara; Napoli, Mariagrazia; Sinicropi, Maria Stefania; Botta, Antonio; Sala, Marina; Carcereri de Prati, Alessandra; Novellino, Ettore; Suzuki, HisanoriEuropean Journal of Medicinal Chemistry (2013),The signaling pathway of Janus tyrosine Kinases-Signal Transducers and Activators of Transcription (JAK-STAT) is activated by a no. of cytokines, hormones (GH, erythropoietin and prolactin), and growth factors. JAK-STAT signaling is involved in regulation of cell proliferation, differentiation and apoptosis. These activities are due to different members of JAK-STAT family consisting of: JAK1, JAK2, JAK3, Tyk2 and STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b, STAT6. Recent studies suggest a key role for STAT family proteins, in particular for STAT3, in selectively inducing and maintaining a pro-carcinogenic inflammatory microenvironment, that promote tumor cells transformation. Moreover, a striking correlation between cancer development/progression and STAT3 persistent activation exists, probably due to STAT3 promoting of the pro-oncogenic inflammatory pathways, like NF-kB, IL-6 and JAK family kinases. Recent study demonstrated that carbazoles can inhibit STAT3 mediated transcription. From these evidences, STAT3 represents a therapeutic target, so we have synthesized a new set of N-alkylcarbazole derivs. substituted in positions 2, 4 and 6, to evaluate their activity on STAT3. Some of these compds. showed an interesting activity as STAT3 selective inhibitors; in particular, compds. 9a 9b and 9c revealed to inhibit the STAT3 activation for the 50%, 90% and 95%, resp. - 29.Timofeeva
, O. A.; Tarasova, N. I.; Zhang, X.; Chasovskikh, S.; Cheema, A. K.; Wong, H.; Brown, M. L.; Dritschilo, A. STAT3 suppresses transcription of pro-apoptotic genes in cancer cells with the involvement of its N-terminal domain Proc. Natl. Acad. Sci. U. S. A. 2013, 110, 1267– 1272[Crossref], [PubMed], [CAS]29.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhvFertb4%253D&md5=08341ca3136e4d2018bd32953eb24927Timofeeva, Olga A.; Tarasova, Nadya I.; Zhang, Xueping; Chasovskikh, Sergey; Cheema, Amrita K.; Wang, Honghe; Brown, Milton L.; Dritschilo, AnatolyProceedings of the National Academy of Sciences of the United States of America (2013),Activation of STAT3 in cancers leads to gene expression promoting cell proliferation and resistance to apoptosis, as well as tumor angiogenesis, invasion, and migration. In the characterization of effects of ST3-H2A2, a selective inhibitor of the STAT3 N-terminal domain (ND), we obsd. that the compd. induced apoptotic death in cancer cells assocd. with robust activation of proapoptotic genes. Using ChIP and tiling human promoter arrays, we found that activation of gene expression in response to ST3-H2A2 is accompanied by altered STAT3 chromatin binding. Using inhibitors of STAT3 phosphorylation and a dominant-neg. STAT3 mutant, we found that the unphosphorylated form of STAT3 binds to regulatory regions of proapoptotic genes and prevents their expression in tumor cells but not normal cells. SiRNA knockdown confirmed the effects of ST3-HA2A on gene expression and chromatin binding to be STAT3 dependent. The STAT3-binding region of the C/EBP-homologous protein (CHOP) promoter was found to be localized in DNaseI hypersensitive site of chromatin in cancer cells but not in nontransformed cells, suggesting that STAT3 binding and suppressive action can be chromatin structure dependent. These data demonstrate a suppressive role for the STAT3 ND in the regulation of proapoptotic gene expression in cancer cells, providing further support for targeting STAT3 ND for cancer therapy. - 30.Timofeeva
, O. A.; Chasovskikh, S.; Lonskaya, I.; Tarasova, N. I.; Khavrutskii, L.; Tarasova, S.; Zhang, X.; Korostyshevskiy, V. R.; Cheema, A.; Zhang, L.; Dakshanamurthy, S.; Brown, M. L.; Dritschilo, A. Mechanisms of unphosphorylated STAT3 transcription factor binding to DNA J. Biol. Chem. 2012, 287, 14192– 14200[Crossref], [PubMed], [CAS]30.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XlvVGlu78%253D&md5=3da0856a85b9d2a51b8797f09c00aeddTimofeeva, Olga A.; Chasovskikh, Sergey; Lonskaya, Irina; Tarasova, Nadya I.; Khavrutskii, Lyuba; Tarasov, Sergey G.; Zhang, Xueping; Korostyshevskiy, Valeriy R.; Cheema, Amrita; Zhang, Lihua; Dakshanamurthy, Sivanesan; Brown, Milton L.; Dritschilo, AnatolyJournal of Biological Chemistry (2012),Phosphorylation of signal transducer and activator of transcription 3 (STAT3) on a single tyrosine residue in response to growth factors, cytokines, interferons, and oncogenes activates its dimerization, translocation to the nucleus, binding to the interferon γ (gamma)-activated sequence (GAS) DNA-binding site and activation of transcription of target genes. STAT3 is constitutively phosphorylated in various cancers and drives gene expression from GAS-contg. promoters to promote tumorigenesis. Recently, roles for unphosphorylated STAT3 (U-STAT3) have been described in response to cytokine stimulation, in cancers, and in maintenance of heterochromatin stability. However, the mechanisms underlying U-STAT3 binding to DNA has not been fully investigated. Here, the authors explore STAT3-DNA interactions by at. force microscopy (AFM) imaging. They obsd. that U-STAT3 mols. bind to the GAS DNA-binding site as dimers and monomers. In addn., U-STAT3 binds to AT-rich DNA sequence sites and recognizes specific DNA structures, such as 4-way junctions and DNA nodes, within neg. supercoiled plasmid DNA. These structures are important for chromatin organization and the data suggest a role for U-STAT3 as a chromatin/genome organizer. Unexpectedly, a C-terminal truncated 67.5-kDa STAT3 isoform recognizes single-stranded spacers within cruciform structures that also have a role in chromatin organization and gene expression. This isoform appears to be abundant in the nuclei of cancer cells and, therefore, may have a role in regulation of gene expression. Taken together, these data highlight novel mechanisms by which U-STAT3 binds to DNA and supports U-STAT3 function as a transcriptional activator and a chromatin/genomic organizer. - 31.Kong
, Y.; Jung, M.; Kang, W.; Grindrod, S.; Velena, A.; Lee, S. A.; Dakshanamurthy, S.; Yang, Y.; Miessau, M.; Zheng, C.; Dritschilo, A.; Brown, M. L. Histone deacetylase cytoplasmic trapping by a novel fluorescent HDAC inhibitor Mol. Cancer Ther. 2011, 10, 1591– 1599[Crossref], [PubMed], [CAS]31.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhtFCjurjK&md5=c8127955235285002cb56c8fdd200b91Kong, Yali; Jung, Mira; Wang, Kan; Grindrod, Scott; Velena, Alfredo; Lee, Sung A.; Dakshanamurthy, Sivanesan; Yang, Yonghong; Miessau, Matthew; Zheng, Chaoyi; Dritschilo, Anatoly; Brown, Milton L.Molecular Cancer Therapeutics (2011),Inhibitors of histone deacetylases (HDAC) are an important emerging class of drugs for the treatment of cancers. HDAC inhibitors are currently under evaluation in clin. trials as single agents and as sensitizers in combinations with chemotherapies and radiation therapy. Although these drugs have important effects on cancer cell growth and functions, the mechanisms underlying HDAC inhibitor activities remain to be fully defined. By using rational drug design, compd. 2, a fluorescent class II HDAC targeting inhibitor, was synthesized and obsd. to accumulate in the cytoplasmic compartments of treated cells, but not in the nuclei. Furthermore, immunostaining of inhibitor exposed cells for HDAC4 showed accumulation of this enzyme in the cytoplasmic compartment with concomitant increased acetylation of tubulin and nuclear histones. These observations support a mechanism by which nuclear histone acetylation is increased as a result of HDAC4 trapping and sequestration in the cytoplasm after binding to compd. 2. The HDAC inhibitor offers potential as a novel theranostic agent, combining diagnostic and therapeutic properties in the same mol. Mol Cancer Ther; 10(9); 1591-9. - 32.Kong
, H. S.; Tian, S.; Kong, Y.; Du, G.; Zhang, L.; Jung, M.; Dritschilo, A.; Brown, M. L. Preclinical studies of YK-4–272, an inhibitor of class III histone deacetylases by disruption of nucleocytoplasmic shuttling Pharm. Res. 2012, 29, 3373– 3383[Crossref], [PubMed], [CAS]32.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhtFShtL7K&md5=ed922486264c800338fdd404a61b88e1Kong, Hye-Sik; Tian, Shuo; Kong, Yali; Du, Guanhua; Zhang, Li; Jung, Mira; Dritschilo, Anatoly; Brown, Milton L.Pharmaceutical Research (2012),Purpose: The HDAC shuttling inhibitor, YK-4-272 functions by restricting nuclear shuttling of Class II HDACs. Pre-clin. investigations of YK-4-272 bioavailability, pharmacokinetics, in vivo toxicity and tumor growth inhibition were performed to det. its potential as an HDAC shuttling disruptor for use in clin. applications. Methods: The soly., lipophilicity, in vitro metabolic stability, in vitro intestinal permeability, and in vivo pharmacokinetics of YK-4-272 were detd. by HPLC methods. The antitumor activity of YK-4-272 was detd. by monitoring athymic Balb/c nude mice bearing PC-3 xenografts. Results: Oral bioavailability of YK-4-272 is supported by its soly. (0.537 mg/mL) and apparent partition coeff. of 2.0. The compd. was chem. and metabolically stable and not a substrate for CYP450. In Caco-2 cell transport studies, YK-4-272 was highly permeable. The time-concn. profile of YK-4-272 in plasma resulted in a Cmax of 2.47 μg/mL at 0.25 h with a AUC of 3.304 μg × h/mL. Treatment of PC-3 tumor xenografts with YK-4-272 showed significant growth delay. Conclusions: YK-4-272 is stable and bio-available following oral administration. Growth inhibition of cancer cells and tumors was obsd. These studies support advancing YK-4-272 for further evaluation as a novel HDAC shuttling inhibitor for use in cancer treatment. - 33.Walls
, T. H.; Grindrod, S.; Beraud, D.; Zhang, L.; Baheti, A. R.; Dakshanamurthy, S.; Patel, M. K.; Brown, M. L.; Macarthur, L. H. Synthesis and biological evaluation of a fluorescent analog of phenytoin as a potential inhibitor of neuropathic pain and imaging agent Bioorg. Med. Chem. 2012, 20, 5269– 5276[Crossref], [PubMed], [CAS]33.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhtFGitrbL&md5=e2ed66fd2258a46c298c6085d37b2d4bWalls, Thomas H.; Grindrod, Scott C.; Beraud, Dawn; Zhang, Li; Baheti, Aparna R.; Dakshanamurthy, Sivanesan; Patel, Manoj K.; Brown, Milton L.; MacArthur, Linda H.Bioorganic & Medicinal Chemistry (2012),Here we report on a novel fluorescent analog of phenytoin as a potential inhibitor of neuropathic pain with potential use as an imaging agent. Compd. 2 incorporated a heptyl side chain and dansyl moiety onto the parent compd. phenytoin and produced greater displacement of BTX from sodium channels and greater functional blockade with greatly reduced toxicity. Compd. 2 reduced mechano-allodynia in a rat model of neuropathic pain and was visualized ex vivo in sensory neuron axons with two-photon microscopy. These results suggest a promising strategy for developing novel sodium channel inhibitors with imaging capabilities. - 34.Sheikh
, K. D.; Banerjee, P. P.; Jagadeesh, S.; Grindord, S. C.; Zhang, L.; Paige, M.; Brown, M. L. Fluorescent epigenetic small molecule induces expression of the tumor suppressor ras-association domain family 1A and inhibits human prostate xenograft J. Med. Chem. 2010, 53, 2376– 2382[ACS Full Text], [CAS]34.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXisVajs7g%253D&md5=7e6d187a13259fb2972b5c3505af4a9eSheikh, Kathryn D.; Banerjee, Partha P.; Jagadeesh, Shankar; Grindrod, Scott C.; Zhang, Li; Paige, Mikell; Brown, Milton L.Journal of Medicinal Chemistry (2010),Epigenetic silencing of Ras-assocn. domain family 1A (RASSF1A) protein in cancer cells results in a disruption of cell cycle control, genetic instability, enhanced cell motility, and apoptotic resistance. Ectopic expression of RASSF1A reverses this tumorigenic phenotype. Thus, small mols. with the ability to restore RASSF1A expression may represent a new class of therapeutic agents. Recently, the authors designed and synthesized a fluorescent carbazole analog of mahanine (alkaloid from Murraya koenigii) that restored RASSF1A mRNA expression. The authors fluorescent lead compd. up-regulated RASSF1A in vitro, potently inhibited human prostate cancer cell proliferation, and fluoresced at a visible wavelength, allowing for the observation of intracellular distribution. The small mol. lead was not acutely toxic ≤550 mg/kg, and dosing at 10 mg/kg reduced human xenograft tumor vol. by about 40%. - 35.Miyaura
, N.; Suzuki, A. Palladium-catalyzed cross-coupling reaction of organoboron compounds Chem. Rev. 1995, 95, 2457– 2483[ACS Full Text], [CAS]35.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2MXoslGiurg%253D&md5=d127b414a75161652876eebc3ed0c486Miyaura, Norio; Suzuki, AkiraChemical Reviews (Washington, D. C.) (1995),A review with >250 refs. including title reactions and their mechanisms, prepn. of organoboron reagents, alkoxycarbonylation and dimerization. - 36.Freeman
, A. W.; Urvoy, M.; Criswell, M. E. Triphenylphosphine-mediated reductive cyclization of 2-nitrobiphenyls: a practical and convenient synthesis of carbazoles J. Org. Chem. 2005, 70, 5014– 5019[ACS Full Text], [CAS]36.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXkt1Wru7s%253D&md5=94995b2ecb6e662cbbf0a0fa1f3ae1efFreeman, Adam W.; Urvoy, Marie; Criswell, Megan E.Journal of Organic Chemistry (2005),The synthesis of a series of substituted carbazoles, e.g., I, from the corresponding 2-nitrobiphenyl derivs. using a novel modification of the Cadogan reaction is described. Cyclization of the 2-nitrobiphenyls was achieved via reductive deoxygenation of the nitro groups using a slight excess of triphenylphosphine in a suitable solvent. A temp. dependence on the extent of conversion under these conditions was obsd., with higher boiling solvents affording higher yields across a range of substrates. The new reaction conditions are very straightforward and convenient to execute, tolerate a broad range of functional groups, and yield carbazole products in the absence of unwanted side products. - 37.Naffziger
, M. R.; Ashburn, B. O.; Perkins, J. R.; Carter, R. G. Diels–Alder approach for the construction of halogenated, o-nitro biaryl templates and application to the total synthesis of the anti-HIV agent siamenol J. Org. Chem. 2007, 72, 9857– 9865[ACS Full Text], [CAS]37.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXns1Ohtrk%253D&md5=c132fbe1f7a1a8242b922be89b21bc00Naffziger, Michael R.; Ashburn, Bradley O.; Perkins, Johanna R.; Carter, Rich G.Journal of Organic Chemistry (2007),A rapid Diels-Alder approach to halogenated biaryl templates is described. These biaryl templates are available in two steps from the corresponding arom. aldehydes. The scope of subsequent Suzuki couplings on the biaryl chlorides is explored. Good tolerance for both electron-donating and electron-withdrawing groups in the coupling process can be achieved. Further functionalization of the biaryl templates is described. Hydrogenation of the nitro moiety with concomitant removal of the benzyl ether yields the o-anilino, o-phenolic polyaryls. Selective redn. of the nitro group can be accomplished. Alternatively, the benzyl ether can be selectively removed under Lewis acidic conditions. The utilization of the Diels-Alder adducts for the synthesis of a series of chlorinated carbazoles via the Cadogan cyclization is also demonstrated. Finally, application of this technol. to the total synthesis of siamenol (I), an anti-HIV agent, is reported. - 38.Nakahara
, K.; Trakoontivakorn, G.; Alzoreky, N. S.; Onishi-Kameyama, M.; Yoshida, M. Antimutagenicity of some edible Thai plants, and a bioactive carbazole alkioid, mahanine, isolated from Micromelum minutum J. Agric. Food Chem. 2002, 50, 4798– 4802 - 39.Roy
, M. K.; Thalang, V. N.; Trakoontivakorn, G.; Nakahara, K. Mahanine, a carbazole alkaloid from Micromelum minutum, inhibits cell growth and induces apotosis through a mitochondrial dependent pathway Br. J. Pharmacol. 2005, 145, 145– 155[Crossref], [PubMed], [CAS]39.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXkt1Shtbk%253D&md5=642aa4f0d8482c8d17e307326e20a812Roy, Molay Kumar; Thalang, Vipaporn Na; Trakoontivakorn, Gassinee; Nakahara, KazuhikoBritish Journal of Pharmacology (2005),Mahanine, a naturally occurring carbazole alkaloid in some Asian vegetables, has been shown to exhibit antimutagenicity, antimicrobial activity, cytotoxicity, and other biol. properties. In the present study, we investigated the effect of mahanine on the activation of the apoptotic pathway in human leukemia U937 cells. Various end points were used to screen for apoptosis: Morphol. changes in cells, the relative nos. of viable and apoptotic cells; translocation of membrane bound phosphatidylserine and DNA anal. We found that mahanine-induced apoptosis in U937 cells involved activation of caspases, including caspase-3, release of cytochrome c into cytosol, loss of mitochondrial membrane permeability, and decreased levels of cellular ATP. Pretreatment of cells with cyclosporine A, prior to/concomitant with exposure to mahanine, effectively prevented the deleterious effects of the alkaloid on cellular integrity and viability. As mitochondrial permeability is known to be important in the regulation of cytochrome c release, our observations indicate that mitochondria are the principal target of mahanine. More specifically, we propose that mahanine causes the mitochondrial membranes to lose their permeability, resulting in caspase-3 activation and apoptosis. - 40.Bromberg
, J. F.; Wrzeszcznska, M. H.; Devgan, G.; Zhao, Y.; Pestell, R. G.; Albanese, C.; Darnell, J. E., Jr. Stat3 as an oncogene Cell 1999, 98, 295– 303[Crossref], [PubMed], [CAS]40.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK1MXlt1Cgt7s%253D&md5=f2e69e246bbfae15a3c5ee7b94340300Bromberg, Jacqueline F.; Wrzeszczynska, Melissa H.; Devgan, Geeta; Zhao, Yanxiang; Pestell, Richard G.; Albanese, Chris; Darnell, James E., Jr.Cell (Cambridge, Massachusetts) (1999),STATs are latent transcription factors that mediate cytokine- and growth factor-directed transcription. In many human cancers and transformed cell lines, Stat3 is persistently activated, and in cell culture, active Stat3 is either required for transformation, enhances transformation, or blocks apoptosis. Here, the substitution of two cysteine residues within the C-terminal loop of the SH2 domain of Stat3 produced a mol. that dimerizes spontaneously, binds to DNA, and activates transcription. The Stat3-C mol. in immortalized fibroblasts causes cellular transformation scored by colony formation in soft agar and tumor formation in nude mice. Thus, the activated Stat3 mol. by itself can mediate cellular transformation and the expts. focus attention on the importance of constitutive Stat3 activation in human tumors. - 41.Nakajima
, K.; Yamanaka, Y.; Nakae, K.; Kojima, H.; Ichiba, M.; Kiuchi, N.; Fukada, T.; Hibi, M.; Hirano, T. A central role for Stat3 in IL-6-induced regulation of growth and differentiation in M1 leukemia cells EMBO J. 1996, 15, 3651– 3658[PubMed], [CAS]41.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK28Xksl2mtbY%253D&md5=7f98221695b15e132b6c11b231b76e5eNakajima, Koichi; Yamanaka, Yojiro; Nakae, Kazuto; Kojima, Hirotada; Ichiba, Makoto; Kiuchi, Nobuo; Kitaoka, Teruichiro; Fukada, Toshiyuki; Hibi, Masahiko; Hirano, ToshioEMBO Journal (1996),Interleukin-6 (IL-6) induces either differentiation or growth of a variety of cells. Little is known about the mol. basis of this cellular decision. The family of signal transducer and activator of transcription (Stat) proteins are involved in signaling through a variety of cytokine and growth factor receptors, although their biol. roles have not been established. To address whether Stat proteins play roles in IL-6-induced growth or differentiation, we introduced two types of mutant Stat3 acting in a dominant-neg. manner into M1 leukemic cells which respond to IL-6 with growth arrest and terminal differentiation. We show that dominant-neg. forms of Stat3 inhibited both IL-6-induced growth arrest at G0/G1 and macrophage differentiation in the M1 transformants. Blocking of Stat activation resulted in inhibition of IL-6-induced repression of c-myb and c-myc. Furthermore, IL-6 enhanced the growth of M1 cells primarily through shortening the length of the G1 period when Stat3 was suppressed. Thus IL-6 generates both growth-enhancing signals and growth arrest- and differentiation-inducing signals at the same time. Stat3 may be a key mol. which dets. the cellular decision from cell growth to differentiation in M1 cells. - 42.Hedvat
, M.; Huszar, D.; Herrmann, A.; Gozgit, J. M.; Schroeder, A.; Sheehy, A.; Buettner, R.; Proia, D.; Kowolik, C. M.; Hix, H.; Armstrong, B.; Bebernitz, G.; Weng, S.; Wang, L.; Ye, M.; McEachern, K.; Morosini, D.; Bell, K.; Alimzhanov, M.; Ioannidis, S.; McCoon, P.; Cao, Z. A.; Yu, H.; Jove, R.; Zinda, M. The JAK2 inhibitor AZD1480 potently blocks Stat3 signaling and oncogenesis in solid tumors Cancer Cell 2009, 16, 487– 497[Crossref], [PubMed], [CAS]42.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXht1eqsbg%253D&md5=9331944af1ea5ab014840218aa928e12Hedvat, Michael; Huszar, Dennis; Herrmann, Andreas; Gozgit, Joseph M.; Schroeder, Anne; Sheehy, Adam; Buettner, Ralf; Proia, David; Kowolik, Claudia M.; Xin, Hong; Armstrong, Brian; Bebernitz, Geraldine; Weng, Shaobu; Wang, Lin; Ye, Minwei; McEachern, Kristen; Chen, Huawei; Morosini, Deborah; Bell, Kirsten; Alimzhanov, Marat; Ioannidis, Stephanos; McCoon, Patricia; Cao, Zhu A.; Yu, Hua; Jove, Richard; Zinda, MichaelCancer Cell (2009),Persistent activation of Stat3 is oncogenic and is prevalent in a wide variety of human cancers. Chronic cytokine stimulation is assocd. with Stat3 activation in some tumors, implicating cytokine receptor-assocd. Jak family kinases. Using Jak2 inhibitors, we demonstrate a central role of Jaks in modulating basal and cytokine-induced Stat3 activation in human solid tumor cell lines. Inhibition of Jak2 activity is assocd. with abrogation of Stat3 nuclear translocation and tumorigenesis. The Jak2 inhibitor AZD1480 suppresses the growth of human solid tumor xenografts harboring persistent Stat3 activity. We demonstrate the essential role of Stat3 downstream of Jaks by inhibition of tumor growth using short hairpin RNA targeting Stat3. Our data support a key role of Jak kinase activity in Stat3-dependent tumorigenesis. - 43.Manshouri
, T.; Quintás-Cardama, A.; Nussenzveig, R. H.; Gaikwad, A.; Estrov, Z.; Prchal, J.; Cortes, J. E.; Kantarjian, H. M.; Verstovsek, S. The JAK kinase inhibitor CP-690, 550 suppresses the growth of human polycythemia vera cells carrying the JAK2V617F mutation Cancer Sci. 2008, 99, 1265– 1273[Crossref], [PubMed], [CAS]43.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1cXnsVOrsbo%253D&md5=85b23c76a0ead47180b7fee84ad061a2Manshouri, Taghi; Quintas-Cardama, Alfonso; Nussenzveig, Roberto H.; Gaikwad, Amos; Estrov, Zeev; Prchal, Josef; Cortes, Jorge E.; Kantarjian, Hagop M.; Verstovsek, SrdanCancer Science (2008),The somatic activating janus kinase 2 mutation (JAK2)V617F is detectable in most patients with polycythemia vera (PV). Here we report that CP-690,550 exerts greater antiproliferative and pro-apoptotic activity against cells harboring JAK2V617F compared with JAK2WT. CP-690,550 treatment of murine factor-dependent cell Patersen-erythropoietin receptor (FDCP-EpoR) cells harboring human wild-type or V617F JAK2 resulted in inhibition of cell proliferation with a 50% inhibitory concn. (IC50) of 2.1 μM and 0.25 μM, resp. Moreover, CP-690,550 induced a significant pro-apoptotic effect on murine FDCP-EpoR cells carrying JAK2V617F, whereas a lesser effect was obsd. for cells carrying wild-type JAK2. This activity was coupled with inhibition of phosphorylation of the key JAK2V617F-dependent downstream signaling effectors signal transducer and activator of transcription (STAT)3, STAT5, and v-akt murine thymoma viral oncogene homolog (AKT). Furthermore, CP-690,550 treatment of ex-vivo-expanded erythroid progenitors from JAK2V617F-pos. PV patients resulted in specific, antiproliferative (IC50 = 0.2 μM) and pro-apoptotic activity. In contrast, expanded progenitors from healthy controls were less sensitive to CP-690,550 in proliferation (IC50 > 1.0 μM), and apoptosis assays. The antiproliferative effect on expanded patient progenitors was paralleled by a decrease in JAK2V617F mutant allele frequency, particularly in a patient homozygous for JAK2V617F. Flow cytometric anal. of expanded PV progenitor cells treated with CP-690,550 suggests a possible transition toward a pattern of erythroid differentiation resembling expanded cells from normal healthy controls. - 44.Zhao
, M.; Gao, F. H.; Wang, J. Y.; Liu, F.; Yuan, H. H.; Zhang, W. Y.; Jiang, B. JAK2/STAT3 signaling pathway activation mediates tumor angiogenesis by upregulation of VEGF and bFGF in non-small-cell lung cancer Lung Cancer 2011, 73, 366– 374[Crossref], [PubMed], [CAS]44.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3Mjit1arsA%253D%253D&md5=ca62ebc5ba2937ff72eb7980e3e284faZhao Mei; Gao Feng-Hou; Wang Jiong-Yi; Liu Feng; Yuan Hai-Hua; Zhang Wen-Ying; Jiang BinLung cancer (Amsterdam, Netherlands) (2011),We investigated the clinical significance of Janus Kinase 2/Signal Transducer and Activator of Transcription 3 (JAK2/STAT3), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) during angiogenesis in non-small-cell lung carcinoma. JAK2, phospho-JAK2 (pJAK2), STAT3, and phospho-STAT3 (pSATA3) were observed in 40/68 (58.8%), 39/68 (57.4%), 49/68 (72.1%) and 40/68 (58.8%) of the cases. The high expression levels of molecules involved in the JAK2/STAT3 signaling pathway were associated with a decreased survival rate. Of the total number of cases, 73.5% were positive for VEGF and 80.9% for bFGF. Microvessel density (MVD), as determined by CD34 staining and morphology, was higher in NSCLC samples with high pJAK2 and pSTAT3 expression, and the patients with high MVD had poor survival status. In addition, the expression of pSTAT3 correlated with VEGF (r=0.593) and bFGF (r=0.519) (p<0.05). Inhibiting JAK2 and knocking down STAT3 both suppressed STAT3 activation and reduced the expression of VEGF and bFGF in A549 and NCI-H292 cells, demonstrating that STAT3 activation was associated with VEGF and bFGF expression in the two human lung carcinoma cell lines. Therefore, STAT3 may be a critical molecular target for powerful intervention in NSCLC anti-angiogenesis therapy. - 45.Yin
, Z. J.; Jin, F. G.; Liu, T. G.; Fu, E. Q.; Xie, Y. H.; Sun, R. L. Overexpression of STAT3 potentiates growth, survival, and radioresistance of non-small-cell lung cancer (NSCLC) cells J. Surg. Res. 2011, 171, 675– 683[Crossref], [PubMed], [CAS]45.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhsVOltLfI&md5=e66e986fa88705c6fb218065035b08a9Yin, Zhen-Jie; Jin, Fa-Guang; Liu, Tong-Gang; Fu, En-Qing; Xie, Yong-Hong; Sun, Rui-LinJournal of Surgical Research (2011),Activation of signal transducer and activator of transcription 3 (STAT3) play important roles in tumorigenesis and tumor progression. The overexpression of STAT3 has been found in various malignancies including non-small-cell lung carcinoma (NSCLC). The purpose of this study was to explore the correlation between overexpression of STAT3 gene and growth, survival, and radiosensitivity of NSCLC cells. Subclones using vector-based short hairpin RNA (shRNA) were established. RT-PCR and Western blot assays were performed to detect the expression of STAT3 mRNA and protein in untransfected or stably transfected NSCLC cells. Then, MTT and soft agar colony assays were performed to det. the effect of STAT3 inhibition on in vitro growth of NSCLC cells. Hoechst staining assay was performed to analyze the effect of STAT3 inhibition on apoptosis of NSCLC cells. Addnl., clonogenic survival assays were performed to detect the effect of STAT3 inhibition on in vitro radiosensitivity of NSCLC cells. Finally, to examine the effect of pSUPER-shSTAT3 on proliferation and radiosensitivity in vivo, a s.c. tumor formation assay in nude mice was performed. Results: We successfully established two stable transfected cell lines (A549/shSTAT3 and SK-MES-1/shSTAT3) in which the expression of STAT3 mRNA and protein was down-regulated. Those two stable subclones showed a significantly dramatic redn. in colony-forming ability and proliferation not only in vitro but also in vivo. The apoptotic rates of A549/shSTAT3 and SK-MES-1/shSTAT3 cells increased to 19.2% and 16.4%, resp. Moreover, shRNA-mediated STAT3 inhibition could also significantly enhance radiosensitivity of NSCLC cells both in vitro and in vivo. Together, the overexpression of STAT3 is correlated with growth, survival, and radioresistance of NSCLC cells, and STAT3 might be a mol. therapeutic target for gene therapy or radiosensitization of NSCLC. - 46.Song
, L.; Turkson, J.; Karras, J. G.; Jove, R.; Haura, E. B. Activation of STAT3 by receptor tyrosine kinases and cytokines regulates survival in human non-small-cell carcinoma cells Oncogene 2003, 22, 4150– 4165[Crossref], [PubMed], [CAS]46.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3sXkvFSkur8%253D&md5=4b7bdafbc03f52cd0a64b061ef39f40eSong, Lanxi; Turkson, James; Karras, James G.; Jove, Richard; Haura, Eric B.Oncogene (2003),Overexpression of receptor tyrosine kinases including the epidermal growth factor receptor (EGF-R) as well as nonreceptor tyrosine kinases, such as Src, have been implicated in the formation of human lung cancers. In addn., cytokines like interleukin-6 (IL-6) have been demonstrated to modulate lung cancer cell growth and elevated levels of IL-6 have been shown to be an adverse prognostic factor for patients with lung cancer. Despite a large body of evidence pointing to their potential importance, few direct studies into the role of signal transducers and activators of transcription (STAT) pathways in human lung cancer have been undertaken. Here the authors demonstrate that multiple nonsmall cell lung cancer cell lines demonstrate constitutive Stat3 DNA-binding activity. Stat3 DNA-binding activity is specifically upregulated by the addn. of epidermal growth factor (EGF), IL-6, and hepatocyte-derived growth factor (HGF). Furthermore, the stimulation of Stat3 DNA-binding activity by EGF requires the activity of EGF-R tyrosine kinase as well as Src-kinase, while the upregulation of Stat3 activity by IL-6 or HGF requires only Src-kinase activity. Treatment of A549 lung cancer cells with PD180970 or SU6656, both pharmacol. inhibitors of Src-kinase, resulted in reduced Src and Stat3 activity, cell cycle arrest in G2, and reduced viability of cells accompanied by induction of apoptosis. Treatment of Stat3-pos. A549 and H358 cells with antisense Stat3 oligonucleotides results in complete loss of Stat3 DNA-binding activity and apoptosis, while Stat3-pos. H1299 cells remained healthy. Finally, an adenoviral vector expressing a dominant-neg. Stat3 isoform results in loss of Stat3 DNA-binding activity, apoptosis, and reduced cellular viability. These results demonstrate a role of Stat3 in transducing survival signals downstream of tyrosine kinases such as Src, EGF-R, and c-Met, as well as cytokines such as IL-6, in human nonsmall cell lung cancers. - 47.OECD. Acute oral toxicity-up and down procedure. OECD Guides for Testing of Chemicals; Organization for Economic Cooperation and Development: Washington, DC, 2001; http://ntp.niehs.nih.gov/iccvam/suppdocs/feddocs/oecd/oecd_gl425-508.pdf.There is no corresponding record for this reference.
- 48.Real
, P.; Sierra, A.; De, J. A.; Segovia, J.; Lopez-Vega, J.; Fernandez- Luna, J. Resistance to chemotherapy via Stat3-dependent overexpression of Bcl-2 in metastatic breast cancer cells Oncogene 2002, 21, 7611– 7618[Crossref], [PubMed], [CAS]48.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38XotFWrtb8%253D&md5=73b6aebcd44e0c591551d7c2ee535cb6Real, Pedro J.; Sierra, Angels; de Juan, Ana; Segovia, Jose C.; Lopez-Vega, Jose M.; Fernandez-Luna, Jose L.Oncogene (2002),Disruption of apoptosis may allow metastatic cell survival and confer resistance to chemotherapeutic drugs. We have analyzed the mol. pathways that activate these survival genes in specific sites of metastasis. Estrogen receptor-neg. breast cancer cell line MDA-MB435 and two metastatic sublines derived from lung (435L) and brain (435B) were analyzed for the expression of members of the Bcl-2 family of apoptosis regulators. The levels of Bcl-2 were higher in the metastatic sublines than in parental cells, which correlated with the activation of Stat3, but not with the expression and/or activation of known bcl-2 transcription factors (CREB and WT1). In the brain subline, both expression of Bcl-2 and Stat3 activation were induced by epidermal growth factor and abrogated after treatment with kinase inhibitors specific for epidermal growth factor receptor or Jak2. Furthermore, transfection of 435B with a dominant-neg. Stat3 markedly reduced the expression of Bcl-2 protein, whereas transient expression of a constitutively active Stat3 increased Bcl-2 in parental 435 cells. In addn., blockade of Stat3 activation by treatment with epidermal growth factor receptor and Jak2 kinase inhibitors or transfection with a dominant neg. Stat3, sensitizes 435B cells to chemotherapy-induced apoptosis. Our data suggest that an increased activation of the Stat3-Bcl-2 pathway in estrogen receptor-neg. metastatic breast cancer cell lines confer a survival advantage to these cells and contribute to their chemoresistance. - 49.Wang
, T.; Niu, G.; Kortylewiski, M.; Burdelya, L.; Shain, K.; Zhang, S.; Bhattacharya, R.; Gabrilovin, D.; Heller, R.; Coppola, D.; Dalton, W.; Jove, R.; Pardoll, D.; Yu, H. Regulation of the innate and adaptive immune responses by Stat-3 signaling in tumor cells Nature Med. 2004, 10, 48– 54[Crossref], [PubMed], [CAS]49.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXlsFGn&md5=3903f2f511796c376fe9cefc35b442ddWang, Tianhong; Niu, Guilian; Kortylewski, Marcin; Burdelya, Lyudmila; Shain, Kenneth; Zhang, Shumin; Bhattacharya, Raka; Gabrilovich, Dmitry; Heller, Richard; Coppola, Domenico; Dalton, William; Jove, Richard; Pardoll, Drew; Yu, HuaNature Medicine (New York, NY, United States) (2004),Although tumor progression involves processes such as tissue invasion that can activate inflammatory responses, the immune system largely ignores or tolerates disseminated cancers. The mechanisms that block initiation of immune responses during cancer development are poorly understood. We report here that constitutive activation of Stat-3, a common oncogenic signaling pathway, suppresses tumor expression of proinflammatory mediators. Blocking Stat-3 in tumor cells increases expression of proinflammatory cytokines and chemokines that activate innate immunity and dendritic cells, leading to tumor-specific T-cell responses. In addn., constitutive Stat-3 activity induces prodn. of pleiotropic factors that inhibit dendritic cell functional maturation. Tumor-derived factors inhibit dendritic cell maturation through Stat-3 activation in progenitor cells. Thus, inhibition of antitumor immunity involves a cascade of Stat-3 activation propagating from tumor to dendritic cells. We propose that tumor Stat-3 activity can mediate immune evasion by blocking both the prodn. and sensing of inflammatory signals by multiple components of the immune system. - 50.Kusaba
, T.; Nakayama, T.; Yamazumi, K.; Yakata, Y.; Yoshizaki, A.; Nagayasu, T.; Sekine, I. Expression of p-STAT3 in human colorectal adenocarcinoma and adenoma; correlation with clinicopathological factors J. Clin. Pathol. 2005, 58, 833– 838[Crossref], [PubMed], [CAS]50.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXpslGnu7s%253D&md5=8bbd6b0b0c0e00f5fb199a52fb460fa9Kusaba, T.; Nakayama, T.; Yamazumi, K.; Yakata, Y.; Yoshizaki, A.; Nagayasu, T.; Sekine, I.Journal of Clinical Pathology (2005),The signal transducer and activator of transcription 3 (STAT3) is a key signaling mol. implicated in the regulation of growth and malignant transformation. Constitutive activation of STATS is seen in several tumor derived cell lines, and in a wide variety of human malignancies. To examine the relation between p-STAT3 (activated form of STAT3) expression and clinicopathol. factors in human colorectal adenocarcinoma and adenoma. Immunohistochem. analyses were carried out on tissues from 44 colorectal adenomas and 95 colorectal adenocarcinomas, comprising 18 intramucosal carcinomas and 77 invasive carcinomas. Seventy seven of these 139 samples (55.4%) showed immunoreactivity for p-STAT3. Pos. staining for p-STATS was seen in 69 of the 95 carcinomas. Only eight of the 44 adenomas showed immunopositivity for p-STATS, resulting in a significant difference between total adenocarcinomas and adenomas (p < 0.001). Among the 95 cases of colorectal adenocarcinoma, p-STAT3 immunoreactivity was significantly correlated with the depth of tumor invasion (p < 0.05), venous invasion (p < 0.05), lymph node metastasis (p < 0.05), and increasing stages of the Dukes' classification (p < 0.01). Expression of p-STAT3 was detected by Western blot anal. in two different cultured human colorectal carcinoma cell lines and six colon carcinoma tissue samples obtained at surgery. This is the first study to report a significant correlation of p-STAT3 expression with the depth of tumor invasion. These findings suggest that p-STAT3 expression is an important factor related to carcinogenesis and/or tumor invasion of colorectal adenocarcinoma. - 51.Ma
, X. T.; Wang, S.; Ye, Y. J.; Du, R. Y.; Cui, Z. R.; Somsouk, M. Constitutive activation of Stat3 signaling pathway in human colorectal carcinoma World J. Gastroenterol 2004, 10, 1569– 1573[PubMed], [CAS]51.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXlsFWgsb4%253D&md5=60a4033b402df902add14b5a0034b9a1Ma, Xiang-Tao; Wang, Shan; Ye, Ying-Jiang; Du, Ru-Yu; Cui, Zhi-Rong; Ma, SomsoukWorld Journal of Gastroenterology (2004),AIM: Signal transducers and activators of transcription (STATs) are a family of transcription factors activated in response to cytokines and growth factors. Constitutive activation of Stat3 has been obsd. in a growing no. of tumor-derived cell lines, as well as tumor specimens from human cancers. The purpose of this study was to investigate the expression of p-Stat3, activated form of Stat3, and its downstream mediators including cyclin D1 and Bcl-xL in colorectal carcinoma (CRC), and to explore the possible mechanism of Stat3 signaling pathway in the tumorigenesis of colorectal carcinoma. METHODS: Tissue samples from 45 patients of primary colorectal carcinoma were selected for studying Stat3 signaling pathway protein expression. Western blot anal. was used to measure the expression of p-Stat3, cyclin D1, and Bcl-xL proteins in colorectal carcinomas. Furthermore, the expression patterns of these proteins were analyzed for their distribution at the cellular level by immunohistochem. staining of the tissues. RESULTS: Protein levels of p-Stat3, cyclin D1, and Bcl-xL were increased in colorectal carcinomas compared with adjacent normal mucosae. Elevated levels of p-Stat3 were correlated with the nodal metastasis and the stage. Overexpression of cyclin D1 was assocd. with the nodal metastasis. There was also a significant correlation between the expressions of p-Stat3 and cyclin D1. CONCLUSION: Constitutive activation of Stat3 may play an important role in the tumorigenesis of colorectal carcinoma, and the detailed mechanism of Stat3 signaling pathway in CRC deserves further investigation. - 52.Kim
, E.; Kim, M.; Woo, D.-H.; Shin, Y.; Shin, J.; Chang, N.; Oh, Y. T.; Kim, H.; Rheey, J.; Nakano, I.; Lee, C.; Joo, K. M.; Rich, J. N.; Nam, D.-H.; Lee, J. Phosphorylation of EZH2 activates STAT3 signaling via STAT3 methylation and promotes tumorigenicity of glioblastoma stem-like cells Cancer Cell 2013, 23, 839– 852[Crossref], [PubMed], [CAS]52.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXnvVGrsb4%253D&md5=24ff8c64a4d23467958d7cecfa0f090cKim, Eunhee; Kim, Misuk; Woo, Dong-Hun; Shin, Yongjae; Shin, Jihye; Chang, Nakho; Oh, Young Taek; Kim, Hong; Rheey, Jingeun; Nakano, Ichiro; Lee, Cheolju; Joo, Kyeung Min; Rich, Jeremy N.; Nam, Do-Hyun; Lee, JeongwuCancer Cell (2013),Glioblastoma multiforme (GBM) displays cellular hierarchies harboring a subpopulation of stem-like cells (GSCs). Enhancer of Zeste Homolog 2 (EZH2), the lysine methyltransferase of Polycomb repressive complex 2, mediates transcriptional repression of prodifferentiation genes in both normal and neoplastic stem cells. An oncogenic role of EZH2 as a transcriptional silencer is well established; however, addnl. functions of EZH2 are incompletely understood. Here, we show that EZH2 binds to and methylates STAT3, leading to enhanced STAT3 activity by increased tyrosine phosphorylation of STAT3. The EZH2-STAT3 interaction preferentially occurs in GSCs relative to non-stem bulk tumor cells, and it requires a specific phosphorylation of EZH2. Inhibition of EZH2 reverses the silencing of Polycomb target genes and diminishes STAT3 activity, suggesting therapeutic strategies. - 53.Berishaj
, M.; Gao, S. P.; Ahmed, S.; Leslie, K.; Al-Ahmadie, H.; Gerald, W. L.; Bornmann, W.; Bromberg, J. F. Stat3 is tyrosine-phosphorylated through the interleukin-6/glycoprotein 130/Janus kinase pathway in breast cancer Breast Cancer Res. 2007, 9, R32 - 54.Zhang
, P.; Zhao, Y.; Zhu, X.; Sedwick, D.; Zhang, X.; Wang, Z. Cross-talk between phospho-STAT3 and PLCγ1 plays a critical role in colorectal tumorigenesis Mol. Cancer Res. 2011, 9, 1418– 1428[Crossref], [PubMed], [CAS]54.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhtlWrtbjN&md5=b5410d8f7efd6afef30336568fe9109dZhang, Peng; Zhao, Yiqing; Zhu, Xiaofeng; Sedwick, David; Zhang, Xiaodong; Wang, ZhengheMolecular Cancer Research (2011),Hyperphosphorylation at the Y705 residue of signal transducer and activator of transcription 3 (STAT3) is implicated in tumorigenesis of leukemia and some solid tumors. However, its role in the development of colorectal cancer is not well defined. To rigorously test the impact of this phosphorylation on colorectal tumorigenesis, we engineered a STAT3 Y705F knock-in to interrupt STAT3 activity in HCT116 and RKO colorectal cancer cells. These STAT3 Y705F mutant cells fail to respond to cytokine stimulation and grow slower than parental cells. These mutant cells are also greatly diminished in their abilities to form colonies in culture, to exhibit anchorage-independent growth in soft agar, and to grow as xenografts in nude mice. These observations strongly support the premise that STAT3 Y705 phosphorylation is crucial in colorectal tumorigenesis. Although it is generally believed that STAT3 functions as a transcription factor, recent studies indicate that transcription-independent functions of STAT3 also play an important role in tumorigenesis. We show here that wild-type STAT3, but not STAT3 Y705F mutant protein, assocs. with phospholipase Cγ1 (PLCγ1). PLCγ1 is a central signal transducer of growth factor and cytokine signaling pathways that are involved in tumorigenesis. In STAT3 Y705F mutant colorectal cancer cells, PLCγ1 activity is reduced. Moreover, overexpression of a constitutively active form of PLCγ1 rescues the transformation defect of STAT3 Y705F mutant cells. In aggregate, our study identifies previously unknown cross-talk between STAT3 and the PLCγ signaling pathways that may play a crit. role in colorectal tumorigenesis. Mol Cancer Res; 9(10); 1418-28. - 55.Lin
, L.; Deangelis, S.; Foust, E.; Fuchs, J.; Li, C.; Li, P.-K.; Schwartz, E. B.; Lesinski, G. B.; Benson, D.; Lü, J.; Hoyt, D.; Lin, J. A novel small molecule inhibits STAT3 phosphorylation and DNA binding activity and exhibits potent growth suppressive activity in human cancer cells Mol. Cancer 2010, 9, 217[Crossref], [PubMed], [CAS]55.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3cfhtFKksg%253D%253D&md5=60b9e2111ec36ec96625043347cc2f5eLin Li; Deangelis Stephanie; Foust Elizabeth; Fuchs James; Li Chenglong; Li Pui-Kai; Schwartz Eric B; Lesinski Gregory B; Benson Don; Lu Jiagao; Hoyt Dale; Lin JiayuhMolecular cancer (2010),BACKGROUND: Targeting Signal Transducer and Activator of Transcription 3 (STAT3) signaling is an attractive therapeutic approach for most types of human cancers with constitutively activated STAT3. A novel small molecular STAT3 inhibitor, FLLL32 was specifically designed from dietary agent, curcumin to inhibit constitutive STAT3 signaling in multiple myeloma, glioblastoma, liver cancer, and colorectal cancer cells. RESULTS: FLLL32 was found to be a potent inhibitor of STAT3 phosphorylation, STAT3 DNA binding activity, and the expression of STAT3 downstream target genes in vitro, leading to the inhibition of cell proliferation as well as the induction of Caspase-3 and PARP cleavages in human multiple myeloma, glioblastoma, liver cancer, and colorectal cancer cell lines. However, FLLL32 exhibited little inhibition on some tyrosine kinases containing SH2 or both SH2 and SH3 domains, and other protein and lipid kinases using a kinase profile assay. FLLL32 was also more potent than four previously reported JAK2 and STAT3 inhibitors as well as curcumin to inhibit cell viability in these cancer cells. Furthermore, FLLL32 selectively inhibited the induction of STAT3 phosphorylation by Interleukin-6 but not STAT1 phosphorylation by IFN-γ. CONCLUSION: Our findings indicate that FLLL32 exhibits potent inhibitory activity to STAT3 and has potential for targeting multiple myeloma, glioblastoma, liver cancer, and colorectal cancer cells expressing constitutive STAT3 signaling. - 56.Liu
, A.; Liu, Y.; Jin, Z.; Hu, Q.; Lin, L.; Jou, D.; Yang, J.; Xu, Z.; Wang, H.; Li, C.; Lin, J. XZH-5 inhibits STAT3 phosphorylation and enhances the cytotoxicity of chemotherapeutic drugs in human breast and pancreatic cancer cells PLoS One 2012, 7, e46624[Crossref], [PubMed], [CAS]56.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsFWgtr7P&md5=7d9b4528b0e47bfbeace38c9c437bb96Liu, Aiguo; Liu, Yan; Jin, Zhigang; Hu, Qun; Lin, Li; Jou, David; Yang, Jing; Xu, Zhenghu; Wang, Hong; Li, Chenglong; Lin, JiayuhPLoS One (2012),Constitutive activation of Signal Transducers and Activators of Transcription 3 (STAT3) signaling is frequently detected in breast and pancreatic cancer. Inhibiting constitutive STAT3 signaling represents a promising mol. target for therapeutic approach. Using structure-based design, we developed a non-peptide cell-permeable, small mol., termed as XZH-5, which targeted STAT3 phosphorylation. XZH-5 was found to inhibit STAT3 phosphorylation (Tyr705) and induce apoptosis in human breast and pancreatic cancer cell lines expressing elevated levels of phosphorylated STAT3. XZH-5 could also inhibit interleukin-6-induced STAT3 phosphorylation in cancer cell lines expressing low phosphorylated STAT3. Inhibition of STAT3 signaling by XZH-5 was confirmed by the down-regulation of downstream targets of STAT3, such as Cyclin D1, Bcl-2, and Survivin at mRNA level. In addn., XZH-5 inhibited colony formation, cell migration, and enhanced the cytotoxicity of chemotherapeutic drugs when combined with Doxorubicin or Gemcitabine. Our results indicate that XZH-5 may be a potential therapeutic agent for breast and pancreatic cancers with constitutive STAT3 signaling. - 57.Gu
, F.-M.; Li, Q.-L.; Gao, Q.; Jiang, J.-H.; Huang, X.-Y.; Pan, J.-F.; Fan, J.; Zhou, J. Sorafenib inhibits growth and metastasis of hepatocellular carcinoma by blocking STAT3 World J. Gastroenterol. 2011, 17, 3922– 3932[Crossref], [PubMed], [CAS]57.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhsVyqtLjI&md5=1a8b2c86486a41c0ffebff5bb889291aGu, Fang-Ming; Li, Quan-Lin; Gao, Qiang; Jiang, Jia-Hao; Huang, Xiao-Yong; Pan, Jin-Feng; Fan, Jia; Zhou, JianWorld Journal of Gastroenterology (2011),Aim: To investigate the inhibitory role and the underlying mechanisms of sorafenib on signal transducer and activator of transcription 3 (STAT3) activity in hepatocellular carcinoma (HCC). Methods: Human and rat HCC cell lines were treated with sorafenib. Proliferation and STAT3 dephosphorylation were assessed. Potential mol. mechanisms of STAT3 pathway inhibition by sorafenib were evaluated. In vivo antitumor action and STAT3 inhibition were investigated in an immunocompetent orthotopic rat HCC model. Results: Sorafenib decreased STAT3 phosphorylation at the tyrosine and serine residues (Y705 and S727), but did not affect Janus kinase 2 (JAK2) and phosphatase shatterproof 2 (SHP2), which is assocd. with growth inhibition in HCC cells. Dephosphorylation of S727 was assocd. with attenuated extracellular signal-regulated kinase (ERK) phosphorylation, similar to the effects of a mitogen-activated protein kinase (MEK) inhibitor U0126, suggesting that sorafenib induced S727 dephosphorylation by inhibiting MEK/ERK signaling. Meanwhile, sorafenib could also inhibit Akt phosphorylation, and both the phosphatidylinositol-3-kinase (PI3K) inhibitor LY294002 and Akt knockdown resulted in Y705 dephosphorylation, indicating that Y705 dephosphorylation by sorafenib was mediated by inhibiting the PI3K/Akt pathway. Finally, in the rat HCC model, sorafenib significantly inhibited STAT3 activity, reducing tumor growth and metastasis. Conclusion: Sorafenib inhibits growth and metastasis of HCC in part by blocking the MEK/ERK/STAT3 and PI3K/Akt/STAT3 signaling pathways, but independent of JAK2 and SHP2 activation. - 58.Hartman
, Z. C.; Poage, G. M.; den Hollander, P.; Tsimelzon, A.; Hill, J.; Panupinthu, N.; Zhang, Y.; Mazumdar, A.; Hilsenbeck, S. G.; Mills, G. B.; Brown, P. H. Growth of triple-negative breast cancer cells relies upon coordinate autocrine expression of the proinflammatory cytokines IL-6 and IL-8 Cancer Res. 2013, 73, 3470– 3480[Crossref], [PubMed], [CAS]58.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXosFSqt7Y%253D&md5=c6e62490546605629e2d6eb80d485812Hartman, Zachary C.; Poage, Graham M.; den Hollander, Petra; Tsimelzon, Anna; Hill, Jamal; Panupinthu, Nattapon; Zhang, Yun; Mazumdar, Abhijit; Hilsenbeck, Susan G.; Mills, Gordon B.; Brown, Powel H.Cancer Research (2013),Triple-neg. breast cancers (TNBC) are aggressive with no effective targeted therapies. A combined database anal. identified 32 inflammation-related genes differentially expressed in TNBCs and 10 proved crit. for anchorage-independent growth. In TNBC cells, an LPA-LPAR2-EZH2 NF-κB signaling cascade was essential for expression of interleukin (IL)-6, IL-8, and CXCL1. Concurrent inhibition of IL-6 and IL-8 expression dramatically inhibited colony formation and cell survival in vitro and stanched tumor engraftment and growth in vivo. A Cox multivariable anal. of patient specimens revealed that IL-6 and IL-8 expression predicted patient survival times. Together these findings offer a rationale for dual inhibition of IL-6/IL-8 signaling as a therapeutic strategy to improve outcomes for patients with TNBCs. - 59.Han
, Y.; Amin, H. M.; Franko, B.; Frantz, C.; Shi, X.; Lai, R. Loss of SHP1 enhances JAK3/STAT3 signaling and decreases proteosome degradation of JAK3 and NPM-ALK in ALK+ anaplastic large-cell lymphoma Blood 2006, 108, 2796– 2803[Crossref], [PubMed], [CAS]59.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28XhtFWgtLzF&md5=170ddd55ede6d0362f421fdc1b68c06aHan, Yajun; Amin, Hesham M.; Franko, Bevin; Frantz, Christine; Shi, Xinzhe; Lai, RaymondBlood (2006),Previous studies showed that most cases of ALK+ anaplastic large-cell lymphoma (ALK+ALCL) do not express SHP1, a tyrosine phosphatase and an important neg. regulator for cellular signaling pathways such as that of JAK/STAT. To fully assess the biol. significance of loss of SHP1 in ALK+ALCL, we transfected SHP1 plasmids into 2 SHP1-, ALK+ALCL cell lines, Karpas 299 and SU-DHL-1. After 24 h of transfection, pJAK3 and pSTAT3 were decreased, and these changes correlated with downregulation of STAT3 downstream targets including cyclin D3, mcl-1, and bcl-2. Expression of SHP1 in these 2 cell lines also resulted in marked decreases in the protein levels of JAK3 and NPM-ALK, and these effects were reversible by proteasome inhibitor MG132. Conversely, when SHP1 expression in SUP-M2 (a SHP1+ ALK+ALCL cell line) was inhibited using siRNA, pSTAT3, pJAK3, JAK3, and NPM-ALK were all up-regulated. Co-immunopptn. studies showed that SHP1 was phys. assocd. with JAK3 and NPM-ALK. SHP1 expression in Karpas 299 and SU-DHL-1 led to significant G1 cell cycle arrest but not apoptosis. To conclude, loss of SHP1 contributes to the pathogenesis of ALK+ALCL by 2 mechanisms: (1) it leaves the tyrosine phosphorylation and activation of JAK3/STAT3 unchecked and (2) it decreases proteasome degrdn. of JAK3 and NPM-ALK. - 60.Zhang
, Q.; Wang, H. Y.; Marzec, M.; Raghunath, P. N.; Nagasawa, T.; Wasik, M. A. STAT3- and DNA methylatransferase 1-mediated epigenetic silencing of SHP-1 tyrosine phosphatase tumor suppressor gene in malignant T lymphocytes Proc. Natl. Acad. Sci. U. S. A. 2005, 102, 6948– 6953[Crossref], [PubMed], [CAS]60.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXksVKnsro%253D&md5=39761f8c9a0732604ea14def8d589ad7Zhang, Qian; Wang, Hong Y.; Marzec, Michal; Raghunath, Puthiyaveettil N.; Nagasawa, Tomohiko; Wasik, Mariusz A.Proceedings of the National Academy of Sciences of the United States of America (2005),Expression of SHP-1 phosphatase, a key neg. regulator of cell signaling, is lost in T cell lymphomas and other malignancies due to DNA methylation of the SHP-1 promoter by a currently undefined mechanism. The authors demonstrate that malignant T cells express DNA methyltransferase (DNMT) 1 and that constantly activated signal transducer and activator of transcription (STAT) 3 is capable of binding in vitro to DNA oligonucleotides corresponding to 4 STAT3 SIE/GAS binding sites identified in the SHP-1 promoter. STAT3, DNMT1, and histone deacetylase 1 form complexes and bind to the SHP-1 promoter in vivo. Treatment with pharmacol. grade DNMT1 anti-sense oligonucleotides and STAT3 small-interfering RNA induces in the malignant T cells DNA demethylation and expression of SHP-1 gene. These data indicate that STAT3 may, in part, transform cells by inducing epigenetic silencing of SHP-1 in cooperation with DNMT1 and, apparently, histone deacetylase 1. Reversal of such gene silencing represents an attractive aim for novel anticancer therapies. - 61.Kang
, H. J.; Yi, Y. W.; Kim, H. J.; Hong, Y. B.; Seong, Y. S.; Bae, I. BRCA1 negatively regulated IGF-1 expression through an estrogen-responsive element-like site Cell Death Dis. 2012, 3, e336[Crossref], [CAS]61.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhtlGgurvN&md5=72a8626c936754a241ab30cb2d412c74Kang, H. J.; Yi, Y. W.; Kim, H. J.; Hong, Y. B.; Seong, Y. S.; Bae, I.Cell Death & Disease (2012),The insulin-like growth factor-1 receptor (IGF-1R) signaling pathway is crit. for both normal mammary gland development and malignant transformation. It has been reported that the IGF-1 stimulates breast cancer cell proliferation and is upregulated in tumors with BRCA1/2 mutations. We report here that IGF-1 is neg. regulated by BRCA1 at the transcriptional level in human breast cancer cells. BRCA1 knockdown (BRCA1-KD) induces the expression of IGF-1 mRNA in MCF7 ceils in an estrogen receptor α (ERα)-dependent manner. We found that both BRCA1 and ERα bind to the endogenous IGF-1 promoter region contg. an estrogen-responsive element-like (EREL) site. BRCA1-KD does not significantly affect ERα binding on the IGF-1 promoter. Reporter anal. demonstrates that BRCA1 could regulate IGF-1 transcripts via this EREL site. In addn., ELISA revealed that de-repression of IGF-1 transcription by BRCA1-KD increases the level of extracellular IGF-1 protein, and secreted IGF-1 seems to increase the phospho-IGF-1Rβ and activate its downstream signaling pathway. Blocking the IGF-1/IGF-1 R/phosphoinositide 3-kinase (PI3K)/AKT pathway either by a neutralizing antibody or by small-mol. inhibitors preferentially reduces the proliferation of BRCA1-KD cells. Furthermore, the IGF-1-EREL-Luc reporter assay demonstrates that various inhibitors, which can inhibit the IGF-1R pathway, can suppress this reporter activity. These findings suggest that BRCA1 defectiveness keeps turning on IGF-1/PI3K/AKT signaling, which significantly contributes to increase cell survival and proliferation. - 62.Yi
, Y. W.; Hong, W.; Kang, H. J.; Kim, H. J.; Zhao, W.; Wang, A.; Seong, Y. S.; Bae, I. Inhibition of the PI3K/AKT pathway potentiates cytotoxicity of EGFR kinase inhibitors in triple-negative breast cancer cells J. Cell. Mol. Med. 2013, 17, 648– 656[Crossref], [PubMed], [CAS]62.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhtFWqsbvO&md5=c5347eb23301a96f9d239c711fc28a4aYi, Yong Weon; Hong, Wooyoung; Kang, Hyo Jin; Kim, Hee Jeong; Zhao, Wenjing; Wang, Antai; Seong, Yeon-Sun; Bae, InsooJournal of Cellular and Molecular Medicine (2013),Triple-neg. breast cancers (TNBCs) are known to be intrinsically resistant to inhibitors for epidermal growth factor receptor (EGFR). Until now, clin. trials for TNBCs using EGFR inhibitors (EGFRis) as single agents have yielded disappointing results. Here, we report that combinatorial treatment using EGFRis, such as gefitinib or erlotinib, with PI3K/AKT pathway inhibitors (PI3K/AKTis) demonstrated a synergistic, anti-proliferative effect in cell lines of the basal-like (BL) subtype, a subtype of TNBC. Western blot anal. revealed that the gefitinib/PI-103 combination significantly reduced the level of both phospho-AKT and phospho-ERK in two susceptible BL subtype cell lines, SUM149PT and MDA-MB-468, whereas it had little or no effect on the level of phospho-ERK in two non-susceptible cell lines (HS578T and MDA-MB-231) of mesenchymal stem-like (MSL) TNBC subtype. The gefitinib/PI-103 combination also significantly induced caspase-3/7-mediated PARP cleavage and reduced two anti-apoptotic proteins, XIAP and Bcl-2 in the susceptible cell lines. In addn., the level of myeloid cell leukemia 1 (Mcl-1) protein was markedly decreased by gefitinib/PI-103 combination in the BL TNBC cells, but showed no significant change by this combination in MSL subtype cells. These results suggest that pharmacol. inhibition of EGFR used in combination of PI3K/AKTis is a potential therapeutic approach to treat a subtype of TNBCs. - 64.Yi
, Y. W.; Kang, H. J.; Kim, H. J.; Kong, Y.; Brown, M. L.; Bae, I. Targeting mutant p53 by a SIRT1 activator YK-3-237 inhibits the proliferation of triple-negative breast cancer cells Oncotarget 2013, 4, 984– 994[CAS]64.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3sfgtVKksw%253D%253D&md5=6f50f4b7f724dcff60b26d406e059487Yi Yong Weon; Kang Hyo Jin; Kim Hee Jeong; Kong Yali; Brown Milton L; Bae InsooOncotarget (2013),Many types of mutations in tumor suppressor p53 are oncogenic through gain-of-function. Therefore, targeting mutant p53 (mtp53) is a promising therapeutic approach to fight against many types of cancers. We report here a small molecule compound YK-3-237 that reduces acetylation of mtp53 and exhibits anti-proliferative effects toward triple-negative breast cancer (TNBC) cells carrying mtp53. YK-3-237 activates SIRT1 enzyme activities in vitro and deacetylation of both mtp53 in a SIRT1-dependent manner. Deacetylation of mtp53 resulted in depletion of mtp53 protein level and up-regulated the expression of WTp53-target genes, PUMA and NOXA. YK-3-237 also induces PARP-dependent apoptotic cell death and arrests the cell cycle at G2/M phase of mtp53 TNBC cells. Taken together, our data suggest that targeting acetylation of mtp53 is a potential target to treat human cancers. - 65.Schneider
, C. A.; Rasband, W. S.; Eliceiri, K. W. NIH image to imageJ: 25 years of image analysis Nature Methods 2012, 9, 671– 675[Crossref], [PubMed], [CAS]65.https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhtVKntb7P&md5=85ab928cd79f1e2f2351c834c0c600f0Schneider, Caroline A.; Rasband, Wayne S.; Eliceiri, Kevin W.Nature Methods (2012),For the past 25 years NIH Image and ImageJ software have been pioneers as open tools for the anal. of scientific images. We discuss the origins, challenges and solns. of these two programs, and how their history can serve to advise and inform other software projects.
