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Seraj, Mohammed Jabed; Harding, Michael A.; Gildea, John J; Welch, Danny R.; Theodorescu, Dan

doi:10.1023/a:1011819621859

Cited by 114 publications

(32 citation statements)

References 26 publications

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“…Rho GDI-2 is a cytosolic protein participating in the regulation of both the GDP/GTP cycle and the membrane association/dissociation cycle of Rho/ Rac proteins. Rho GDI proteins are in general overexpressed in breast tumors and human bladder cancer and their increased synthesis correlates with malignancy [37,38]. In agreement with our findings, Kovarova et al [39] have reported, by 2-D map analysis followed by MALDI-MS, that Rho GDI proteins are significantly downregulated in CEM T-lymphoblastic leukemia cell lines after treatment with bohemine.…”

Section: Downregulated Proteinssupporting

confidence: 92%

Proteomic analysis of pancreatic ductal carcinoma cells treated with 5‐aza‐2'‐deoxycytidine

Cecconi¹,

Astner²,

Donadelli

et al. 2003

Electrophoresis

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A pancreatic adenocarcinoma cell line (PaCa44), which contains, among other alterations, a methylated p16 promoter, was treated with a chemoterapeutic agent, 5-aza-2'-deoxycytidine (DAC), in order to evaluate the effect of this drug on cell growth and protein expression. Cell proliferation was strongly inhibited by a 24 h DAC treatment and this inhibition lasted for at least 10 days. Master maps of control and treated PaCa44 cells were generated by analysis with the PDQuest software. The comparison between such maps showed up- and downregulation of 45 polypeptide chains, of which 32 were downregulated and 13 upregulated, out of a total of 700 spots detected by a medium-sensitivity stain, micellar Coomassie Brilliant Blue. Fingerprinting by mass spectrometry analysis enabled the identification of 36 of these spots. Among the major changes in DAC-treated cells: cofilin and profilin 1 are silenced; coactosin, peptidyl-propyl cis-trans isomerase A and cystatin B are decreased by 22, 16- and 15-fold, respectively; stress-70 protein, superoxide dismutase and protein disulfide isomerase A3 are increased by 13-, 11-, and 5-fold, respectively. The significance of some of these major changes is discussed.

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Section: Downregulated Proteinssupporting

confidence: 92%

Proteomic analysis of pancreatic ductal carcinoma cells treated with 5‐aza‐2'‐deoxycytidine

Cecconi¹,

Astner²,

Donadelli

et al. 2003

Electrophoresis

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“…It appears, therefore, that rather than constitutive activation, accelerated GDP-GTP exchange may be more important for Rac1-mediated oncogenesis, as is the case for Rac1b. In addition, Seraj et al (2000) have shown that loss of RhoGDI2 expression is associated with invasive and metastatic human cancers. Therefore, the inability of RhoGDI to interact with Rac1b is also consistent with a role for this splice variant in oncogenesis.…”

Section: Discussionmentioning

confidence: 99%

Rac1b, a tumor associated, constitutively active Rac1 splice variant, promotes cellular transformation

et al. 2004

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A novel splice variant of Rac1, designated Rac1b, is expressed in human breast and colon carcinoma cells. Rac1b contains an additional 19 amino-acid insert immediately behind the switch II domain, a region important for Rac1 interaction with regulators and effectors. Recent studies showed that Rac1b exhibited the biochemical properties of a constitutively activated GTPase, yet it showed impaired interaction with downstream effectors, suggesting that Rac1b may be defective in biological activity. Whether Rac1b is a biologically active protein was not addressed. Therefore, we evaluated the biochemical, signaling and growth-promoting properties of authentic Rac1b. Similar to previous observations, we found that Rac1b showed enhanced intrinsic guanine nucleotide exchange activity, impaired intrinsic GTPase activity, and failed to interact with RhoGDI. Surprisingly, we found that Rac1b, like the constitutively-activated and transforming Rac1(Q61L) mutant, promoted growth transformation of NIH3T3 cells. Rac1b-expressing cells also showed a loss of density-dependent and anchorage-dependent growth. Surprisingly, unlike activated Rac1(61L), Rac1b did not show enhanced activation of the nuclear factor jB (NF-jB) transcription factor or stimulate cyclin D1 expression, the signaling activities that best correlate with Rac1 transforming activity. However, Rac1b did promote activation of the AKT serine/threonine kinase. Therefore, we suggest that Rac1b selectively activates a subset of Rac1 downstream signaling pathways to facilitate cellular transformation.

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“…PC-SPES was administered orally (gavage) at a concentration of 250 mg/kg/day as previously described (Kubota et al, 2000). Mice were weighed every week and tumor sizes were measured twice a week as described (Seraj et al, 2000). Mice were euthanized 40 days postinjection.…”

Section: In Vivo Tumor Growth and Statistical Analysismentioning

confidence: 99%

Gene profiling and promoter reporter assays: Novel tools for comparing the biological effects of botanical extracts on human prostate cancer cells and understanding their mechanisms of action

Bigler

Gulding

Dann³

et al. 2003

Oncogene

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The use of botanical mixtures is commonplace in patients with prostate cancer, yet the majority of these products have not been rigorously tested in clinical trials. Here we use PC-SPES, a combination of eight herbs that has been shown to be effective in clinical trials in patients with prostate cancer, as a model system to demonstrate 'proof of principle' as to how gene expression profiling coupled with promoter assays can evaluate the effect of herbal cocktails on human prostate cancer. In addition, we demonstrate how such approaches may be used for standardization of herbal extract activity by comparing the gene profile of PC-SPES with that of PC-CARE, a product with a similar herbal composition. Since prior studies have shown that PC-SPES contains estrogenic organic compounds, and such compounds are known to affect prostate cancer, an important issue is whether these are the primary drivers of the gene profile. Our data suggest that gene expression profiles of LNCaP human prostate cancer cells in response to PC-SPES are different from those found when diethylstilbestrol (DES), a synthetic estrogen, is used, suggesting that the estrogenic moieties within PC-SPES do not drive this expression signature. In contrast, the expression profile of PC-CARE was almost identical to that of DES, highlighting that mixtures containing similar herbal compositions do not necessarily result in similar biological activities. Interestingly, these three agents cause similar in vitro morphological changes and growth effects on LNCaP. To validate the expression profiling data, we evaluated the protein expression and promoter activity of prostate-specific antigen (PSA), a gene induced by PC-SPES but repressed by DES. In order to gain a mechanistic understanding of how PC-SPES and DES affect PSA expression differently, LNCaP cells were transiently transfected with wild-type and mutagenized PSA promoter, ARE concatemers and appropriate controls. We provide evidence that androgen response elements (ARE) II and III within the promoter region are responsible for the suppressive effects of DES and stimulatory effects of PC-SPES. In addition, we show that the effects on PSA transcription are ARE specific in the case of DES while PC-SPES affects this promoter nonspecifically. In conclusion, expression profiling coupled with mechanistic target validation yield valuable clues as to the mode of action of complex botanical mixtures and provides a new way to compare objectively mixtures with similar components either for effect or quality assurance prior to their use in clinical trials.

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Cited by 114 publications

References 26 publications

Proteomic analysis of pancreatic ductal carcinoma cells treated with 5‐aza‐2'‐deoxycytidine

Proteomic analysis of pancreatic ductal carcinoma cells treated with 5‐aza‐2'‐deoxycytidine

Rac1b, a tumor associated, constitutively active Rac1 splice variant, promotes cellular transformation

Gene profiling and promoter reporter assays: Novel tools for comparing the biological effects of botanical extracts on human prostate cancer cells and understanding their mechanisms of action

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