Leiodermatolide, a novel marine natural product, has potent cytotoxic and antimitotic activity against cancer cells, appears to affect microtubule dynamics, and exhibits antitumor activity.

Pancreatic cancer, the fourth leading cause of cancer death in the United States, has a negative prognosis because metastasis occurs before symptoms manifest. Leiodermatolide, a polyketide macrolide with antimitotic activity isolated from a deep water sponge of the genus Leiodermatium, exhibits potent and selective cytotoxicity toward the pancreatic cancer cell lines AsPC-1, PANC-1, BxPC-3, and MIA PaCa-2, and potent cytotoxicity against skin, breast and colon cancer cell lines. Induction of apoptosis by leiodermatolide was confirmed in the AsPC-1, BxPC-3 and MIA PaCa-2 cells. Leiodermatolide induces cell cycle arrest but has no effects on in vitro polymerization or depolymerization of tubulin alone, while it enhances polymerization of tubulin containing microtubule associated proteins (MAPs). Observations through confocal microscopy show that leiodermatolide, at low concentrations, causes minimal effects on polymerization or depolymerization of the microtubule network in interphase cells, but disruption of spindle formation in mitotic cells. At higher concentrations, depolymerization of the microtubule network is observed. Visualization of the growing microtubule in HeLa cells expressing GFP-tagged plus end binding protein EB-1 showed that leiodermatolide stopped the polymerization of tubulin. These results suggest that leiodermatolide may affect tubulin dynamics without directly interacting with tubulin and hint at a unique mechanism of action. In a mouse model of metastatic pancreatic cancer, leiodermatolide exhibited significant tumor reduction when compared to gemcitabine and controls. The antitumor activities of leiodermatolide, as well as the proven utility of antimitotic compounds against cancer, make leiodermatolide an interesting compound with potential chemotherapeutic effects that may merit further research.

Sensitization of pancreatic cancer cells to radiation by cerium oxide nanoparticle-induced ROS production.

Side effect of radiation therapy (RT) remains the most challenging issue for pancreatic cancer treatment. In this report we determined whether and how cerium oxide nanoparticles (CONPs) sensitize pancreatic cancer cells to RT. CONP pretreatment enhanced radiation-induced reactive oxygen species (ROS) production preferentially in acidic cell-free solutions as well as acidic human pancreatic cancer cells. In acidic environments, CONPs favor the scavenging of superoxide radical over the hydroxyl peroxide resulting in accumulation of the latter whereas in neutral pH CONPs scavenge both. CONP treatment prior to RT markedly potentiated the cancer cell apoptosis both in culture and in tumors and the inhibition of the pancreatic tumor growth without harming the normal tissues or host mice. Taken together, these results identify CONPs as a potentially novel RT-sensitizer as well as protectant for improving pancreatic cancer treatment. FROM THE CLINICAL EDITOR: Pancreatic tumors remain some of the most notoriously treatment-unresponsive malignancies. Cerium oxide nanoparticles may be capable of sensitizing such cells to radiotherapy, as demonstrated in this study.

MGMT inhibition restores ERα functional sensitivity to antiestrogen therapy.

Antiestrogen therapy resistance remains a huge stumbling block in the treatment of breast cancer. We have found significant elevation of O(6) methylguanine DNA methyl transferase (MGMT) expression in a small sample of consecutive patients who have failed tamoxifen treatment. Here, we show that tamoxifen resistance is accompanied by upregulation of MGMT. Further we show that administration of the MGMT inhibitor, O(6)-benzylguanine (BG), at nontoxic doses, leads to restoration of a favorable estrogen receptor alpha (ERα) phosphorylation phenotype (high p-ERα Ser167/low p-ERα Ser118), which has been reported to correlate with sensitivity to endocrine therapy and improved survival. We also show BG to be a dual inhibitor of MGMT and ERα. In tamoxifen-resistant breast cancer cells, BG alone or in combination with antiestrogen (tamoxifen [TAM]/ICI 182,780 [fulvestrant, Faslodex]) therapy enhances p53 upregulated modulator of apoptosis (PUMA) expression, cytochrome C release and poly (ADP-ribose) polymerase (PARP) cleavage, all indicative of apoptosis. In addition, BG increases the expression of p21(cip1/waf1). We also show that BG, alone or in combination therapy, curtails the growth of tamoxifen-resistant breast cancer in vitro and in vivo. In tamoxifen-resistant MCF7 breast cancer xenografts, BG alone or in combination treatment causes significant delay in tumor growth. Immunohistochemistry confirms that BG increases p21(cip1/waf1) and p-ERα Ser167 expression and inhibits MGMT, ERα, p-ERα Ser118 and ki-67 expression. Collectively, our results suggest that MGMT inhibition leads to growth inhibition of tamoxifen-resistant breast cancer in vitro and in vivo and resensitizes tamoxifen-resistant breast cancer cells to antiestrogen therapy. These findings suggest that MGMT inhibition may provide a novel therapeutic strategy for overcoming antiestrogen resistance.

Chemopreventive effects of tolfenamic acid against esophageal tumorigenesis in rats.

The primary objective of this study is to identify small molecules that target critical transcription factors for potential application in the chemoprevention of esophageal cancer. Specificity proteins (Sp) play a critical role in the growth and metastasis of several malignancies including esophageal cancer. Researchers at the M. D. Anderson Cancer Center Orlando Cancer Research Institute have reported previously that tolfenamic acid (TA) inhibits cancer cell proliferation and tumor growth through the degradation of Sp1, Sp3, and Sp4. We evaluated the chemopreventive properties of TA against esophageal tumorigenesis in N-nitrosomethylbenzylamine (NMBA)-induced murine tumor model. Fischer-344 rats were treated with NMBA (0.5 mg/kg s.c. 3 times a week) for 5 weeks to initiate the tumor formation, and then treated with 50 mg/kg TA from week 6 through week 25. Tumor incidence, tumor multiplicity (number of papilloma per rat), and tumor volume were evaluated after 25 weeks. All rats in the control group that received only NMBA developed lesions (100% incidence), while the TA-treated group showed significantly lower (33%) tumor incidence and tumor multiplicity. Furthermore, the tumor volume was significantly diminished in the TA-treated group when compared with the control group. Using small molecules such as TA to target key transcription factors associated with tumorigenesis for the prevention of esophageal malignancies is a new and promising strategy. Results of the current study provide evidence that TA, when given orally after tumor initiation, can significantly suppress tumorigenesis induced by carcinogenic nitrosamines in rats. These appealing results demonstrate that TA may potentially serve as an effective chemopreventive agent in patient populations vulnerable to esophageal cancer.

Harnessing nanoparticles to improve toxicity after head and neck radiation.

This article reports the evaluation of cerium oxide (CeO(2)) nanoparticles' ability to decrease xerostomia and radiation-induced dermatitis in mice after head and neck radiation. Mice were irradiated using an IC160 x-ray system. Two cohorts were included: (A) No-radiation and (B) 30 Gy/6 fractions, and were randomized into three groups: (1) saline, (2) 15 nM CeO(2) and (3) 15 µM CeO(2). Stimulated salivary flow and radiation-induced dermatitis were evaluated post radiation. Stimulated sialometry demonstrated improved salivary production in all CeO(2) groups in comparison with controls (flow: 204 vs. 115 µL/10 minutes, P = 0.0002). One week post radiation, G-III dermatitis decreased in the 15 µM group in comparison with controls (10% versus 100% incidence, respectively). There was decreased skin hyperpigmentation at 12 weeks in the 15-µM group in comparison with 15-nM and non-CeO(2) groups (50%, 70%, and 90% G-II, respectively). This study suggests that CeO(2) may be radioprotective for salivary production and reduces G-III dermatitis and skin hyperpigmentation incidence. CeO(2) as radioprotectant may be a feasible concept during radiotherapy. FROM THE CLINICAL EDITOR: This study demonstrates in a mouse model that cerium oxide (CeO(2)) nanoparticles may provide an important mechanism in preventing radiation induced xerostomia, a common complication of head and neck radiation treatments.

Tolfenamic acid decreases c-Met expression through Sp proteins degradation and inhibits lung cancer cells growth and tumor formation in orthotopic mice.

The nonsteroidal anti-inflammatory drug (NSAID), tolfenamic acid (TA) is emerging as a new anti-cancer agent. TA induces the degradation of specific Specificity protein (Sp) transcription factors, Sp1, Sp3 and Sp4 which are associated with tumor growth and metastasis. In this study we have evaluated the effect of TA on lung cancer using both in vitro and in vivo models. TA in a dose dependent manner inhibited proliferation and cell viability of two different lung cancer cells, A549 and CRL5803. TA treatment for 48 h significantly decreased the expression of Sp1, Sp3 and Sp4. The hepatocyte growth factor receptor, c-Met is overexpressed in a variety of cancers including lung cancer and Sp proteins mediate the regulation of c-Met. TA diminished the expression of c-Met protein and modulates its downstream signaling pathway. Furthermore, TA treatment significantly increased the number of apoptotic cells and pro-apoptotic markers c-PARP and Bax confirming the activation of apoptotic pathways. In vivo studies using the orthotopic mice model for lung cancer showed that TA (25 mg/kg/2 days and 50 mg/kg/2 days) resulted in a dose dependent decrease in tumor formation. The immunohistochemical staining of lung tissue showed high expression of Sp1, Sp3, Sp4, c-Met and phospho Met in control group and a dose dependent decrease in TA treated groups. The crucial findings of this study support that targeting c-Met with a potent inhibitor of Sp proteins is a robust strategy for the implications in lung cancer treatment and TA can serve as a therapeutic agent for this devastating disease.

Tolfenamic acid inhibits ovarian cancer cell growth and decreases the expression of c-Met and survivin through suppressing specificity protein transcription factors.

OBJECTIVE: The aberrant expression of hepatocyte growth factor and its receptor c-Met are associated with aggressive disease and poor prognosis in a variety of human malignancies including ovarian cancer (OC). Specificity protein (Sp) transcription factors have high relevance in the signaling cascade associated with c-Met activation. Tolfenamic acid (TA), a NSAID, is known to induce the degradation of Sp proteins, which have been negatively associated with survival in some cancer patients. Our aim was to examine the anti-OC activity of TA using in vitro and in vivo models and asses the inhibitory effects of this novel compound. METHODS: We developed OC sub-cell lines (AF1-3) derived from the original SKOV3 that are more resistant to IFNα-2b and more tumorigenic in nude mice than the original cells. We tested the anti-cancer activity of TA using ovarian cancer cells, SKOV3-AF2 and ES-2. The cells were treated with DMSO (vehicle) or TA (25/50/100 µM) and cell viability was measured at 24, 48, and 72 h. Cell lysates were prepared following 48 h treatment (50µM) and evaluated the expression of Sp proteins (Sp1/Sp3/Sp4), c-Met, survivin, Bcl2, and cleaved polyADP-ribose polymerase (c-PARP) through Western blot analysis. Caspase-3 activity was assessed with Caspase-Glo kit. Cell cycle distribution and apoptosis were analyzed using BD FACSCalibur flow cytometer. For in vivo studies mice were subcutaneously injected with ES-2 cells and treated with vehicle or TA (50 mg/kg/thrice weekly). RESULTS: TA significantly inhibited the growth of SKOV3 (AF1-3) and ES-2 cells. The expression of Sp proteins and c-Met was significantly decreased suggesting that TA could be targeting c-Met through degradation of Sp proteins. TA greatly increased the apoptotic fraction (Annexin V positive), c-PARP expression and caspase-3 activity. TA significantly decreased Bcl2 expression and induced G0/G1 cell cycle arrest. In vivo studies revealed that TA significantly inhibited tumor weight and volume in mice. These results show that TA has a profound inhibitory effect on tumor growth in mice, reduces OC cells proliferation, induces apoptosis, cell cycle arrest and Sp proteins degradation. TA also inhabited the expression of survivin that is associated with radiation resistance and suggests that apart from its tumor suppressant effects, TA can also enhance the tumor response to radiotherapy. CONCLUSIONS: These data clearly show that TA effectively inhibits OC cell growth in vitro and in vivo. This study represents potential role(s) of TA that suppresses OC cell growth, and may enhance tumor response to radiotherapy, and have implications in OC treatment.

A facile nanoparticle immunoassay for cancer biomarker discovery.

BACKGROUND: Gold nanoparticles (AuNPs) scatter light intensely at or near their surface plasmon wavelength region. Using AuNPs coupled with dynamic light scattering (DLS) detection, we developed a facile nanoparticle immunoassay for serum protein biomarker detection and analysis. A serum sample was first mixed with a citrate-protected AuNP solution. Proteins from the serum were adsorbed to the AuNPs to form a protein corona on the nanoparticle surface. An antibody solution was then added to the assay solution to analyze the target proteins of interest that are present in the protein corona. The protein corona formation and the subsequent binding of antibody to the target proteins in the protein corona were detected by DLS. RESULTS: Using this simple assay, we discovered multiple molecular aberrations associated with prostate cancer from both mice and human blood serum samples. From the mice serum study, we observed difference in the size of the protein corona and mouse IgG level between different mice groups (i.e., mice with aggressive or less aggressive prostate cancer, and normal healthy controls). Furthermore, it was found from both the mice model and the human serum sample study that the level of vascular endothelial growth factor (VEGF, a protein that is associated with tumor angiogenesis) adsorbed to the AuNPs is decreased in cancer samples compared to non-cancerous or less malignant cancer samples. CONCLUSION: The molecular aberrations observed from this study may become new biomarkers for prostate cancer detection. The nanoparticle immunoassay reported here can be used as a convenient and general tool to screen and analyze serum proteins and to discover new biomarkers associated with cancer and other human diseases.

Loss of tyrosine phosphatase-dependent inhibition promotes activation of tyrosine kinase c-Src in detached pancreatic cells.

Despite an intense focus on novel therapeutic strategies, pancreatic adenocarcinoma remains one of the deadliest human malignancies. The frequent and rapid mortality associated with pancreatic cancer may be attributed to several factors, including late diagnosis, rapid tumor invasion into surrounding tissues, and formation of distant metastases. Both local invasion and metastasis require disruption of tumor cell contacts with the extracellular matrix. Detachment of normal cells from the extracellular matrix leads to a form of programmed cell death termed anoikis. Pancreatic cancer cells avert anoikis by activation of signaling pathways that allow for adhesion-independent survival. In the present studies, cellular signaling pathways activated in detached pancreatic cancer cells were examined. We demonstrate a rapid and robust activation of Src kinase in detached pancreatic cancer cells, relative to adherent. Src autophosphorylation rapidly returned to baseline levels upon reattachment to tissue culture plastic, in the presence or absence of specific extracellular matrix proteins. Treatment of pancreatic cancer cells with tyrosine phosphatase inhibitors increased steady-state Src autophosphorylation in adherent cells and abrogated the detachment-induced increase in Src autophosphorylation. Src was found to co-immunoprecipitate with the Src homology 2 (SH2) domain containing protein tyrosine phosphatase (SHP-2) in pancreatic cancer cells, suggesting that SHP-2 may participate in regulation of Src autophosphorylation in adherent cells. Src family kinase (SFK) dependent increases in Akt and Jun N-terminal kinase (JNK) phosphorylation were observed in detached cells, indicating the potential for Src-dependent activation of survival and stress pathways in pancreatic cancer cells that have detached from the extracellular matrix.

A label-free nanoparticle aggregation assay for protein complex/aggregate detection and study.

The detection, analysis, and understanding of protein complexes/aggregates and their formation process are extremely important for biomolecular research, diagnosis, and biopharmaceutical development. Unfortunately, techniques that can be used conveniently for protein complex/aggregate detection and analysis are very limited. Using gold nanoparticle immunoprobes coupled with dynamic light scattering (DLS), we developed a label-free nanoparticle aggregation immunoassay (NanoDLSay) for protein aggregate detection and study. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH), a protein target used routinely in Western blot as a loading control, is demonstrated here as an example. Through this study, we discovered that GAPDH has a strong tendency to form large aggregates in certain buffer solutions at a concentration range of 10-25 microg/ml. The strong light scattering property of gold nanoparticles immunoprobes greatly enhanced the sensitivity of the dynamic light scattering for protein complex/aggregate detection. In contrast to fluorescence techniques for protein complex and aggregation study, the protein targets do not need to be labeled with fluorescent probe molecules in NanoDLSay. NanoDLSay is a very convenient and sensitive tool for protein complex/aggregate detection and study.

A metastatic colon cancer model using nonoperative transanal rectal injection.

BACKGROUND: This study aimed to develop a noninvasive orthotopic model for metastasis of colon and rectal cancer using a transanal approach. Currently, the most accurate orthotopic representation of metastatic human colon cancer is via a cecal injection. The transanal model allows for further examination of systemic immune responses, tumor take, and onset of metastasis without prior surgical intervention. METHODS: For this study, 60 Balb/c mice were anesthetized and subjected to gentle anal dilation using blunt-tipped forceps at the anal opening. Murine colon cancer parental CT26 or luciferase-labeled CT26 (CT26-luc) cells were injected submucosally into the distal posterior rectum (30 CT26 and 30 CT26 injections) at concentrations of 2.5 x 10(4), 1 x 10(5), and 1 x 10(6) in a volume of 50 microl. Tumor growth and metastatic development was monitored at 5-day intervals for 50 days. All the remaining mice were killed on postinjection day 50. RESULTS: The optimal concentration for metastasis and survival of the mice was 2.5 x 10(4) cells. Higher concentrations of cells yielded higher mortality but did not result in metastasis. The overall success of tumor growth in both experiments using the transanal rectal injection was 65%. Histology showed that all tumors were poorly differentiated adenocarcinomas. Two mice (3.3%) from the 2.5 x 10(4) CT26-luc group showed metastatic colonic adenocarcinoma to the liver on postinjection day 50. CONCLUSION: Transanal rectal injection of colon cancer cells offers a nonoperative orthotopic murine model for colon cancer that may lead to the development of metastasis. By using an orthotopic model, more aspects of metastatic colon cancer can be evaluated without the influence of a previous abdominal incision. These results warrant more investigation.

Cerium oxide nanoparticles protect gastrointestinal epithelium from radiation-induced damage by reduction of reactive oxygen species and upregulation of superoxide dismutase 2.

The ability of rare earth cerium oxide (CeO(2)) nanoparticles to confer radioprotection against gastrointestinal epithelium was examined. The pretreatment of normal human colon cells (CRL 1541) with varying concentrations of CeO(2) nanoparticles 24 hours before single-dose radiation exposure conferred protection from radiation-induced cell death by reducing the amount of reactive oxygen species produced and increasing the expression of superoxide dismutase 2 (SOD2), in a dose-dependent manner. In subsequent experiments athymic nude mice were pretreated with intraperitoneal injections of CeO(2) nanoparticles before a single dose of radiation to the abdominal area. Immunohistochemical analysis show a decrease in TUNEL- and caspase 3-positive cells in the colonic crypt, 4 hours after radiation. In sharp contrast, a significant increase in SOD2 expression was observed. In the end, these studies suggest that CeO(2) nanoparticles protect the gastrointestinal epithelium against radiation-induced damage by (1) acting as free-radical scavengers and (2) increasing the production of SOD2 before radiation insult. FROM THE CLINICAL EDITOR: In this study, the ability of rare earth cerium oxide (CeO(2)) nanoparticles to confer radioprotection was examined. The results suggest that CeO(2) nanoparticles protect the gastrointestinal epithelium against radiation-induced damage both by acting as free-radical scavengers and by increasing the production of SOD2 before radiation insult.

Phosphate ester hydrolysis of biologically relevant molecules by cerium oxide nanoparticles.

In an effort to characterize the interaction of cerium oxide nanoparticles (CNPs) in biological systems, we explored the reactivity of CNPs with the phosphate ester bonds of p-nitrophenylphosphate (pNPP), ATP, o-phospho-l-tyrosine, and DNA. The activity of the bond cleavage for pNPP at pH 7 is calculated to be 0.860 ± 0.010 nmol p-nitrophenol/min/µg CNPs. Interestingly, when CNPs bind to plasmid DNA, no cleavage products are detected. While cerium(IV) complexes generally exhibit the ability to break phosphorus-oxygen bonds, the reactions we report appear to be dependent on the availability of cerium(III) sites, not cerium(IV) sites. We investigated the dephosphorylation mechanism from the first principles and find the reaction proceeds through inversion of the phosphate group similar to an S(N)2 mechanism. The ability of CNPs to interact with phosphate ester bonds of biologically relevant molecules has important implications for their use as potential therapeutics.

Tolfenamic acid enhances pancreatic cancer cell and tumor response to radiation therapy by inhibiting survivin protein expression.

Survivin is overexpressed in most human cancers, including pancreatic adenocarcinoma. Expression of survivin is regulated by specificity protein (Sp) proteins and related to resistance to radiation therapy. Tolfenamic acid induces Sp protein degradation in several cancer cell lines. The purpose of this study is to investigate whether tolfenamic acid inhibits survivin expression and sensitizes pancreatic cancer cells/tumor to radiotherapy. Panc1 and L3.6pl cells have been used to study the effect of radiation on survivin expression and to investigate the efficacy of tolfenamic acid in enhancing the response to radiation therapy. In addition, an orthotopic model for human pancreatic cancer has been used to confirm the efficacy of tolfenamic acid to enhance tumor response to radiation in vivo. Pancreatic cancer cell lines express variable levels of survivin mRNA/protein, which correlate with their radiosensitivity. Radiation increased survivin promoter activity and protein expression in Panc1 and L3.6pl cells and tolfenamic acid inhibited both constitutive and radiation-induced survivin protein expression and enhanced the response of pancreatic cancer cells to radiation therapy. In vivo studies show that tolfenamic acid enhanced the radiation-induced apoptosis associated with decreased survivin expression in tumors and this correlates with the enhanced response of these tumors to the radiation. Thus, tolfenamic acid significantly enhances pancreatic cancer cells/tumor response to radiation therapy. The underlying mechanism includes tolfenamic acid-induced degradation of Sp proteins, which in tumor decreases expression of the Sp-dependent antiapoptotic protein survivin. These preclinical data suggest that tolfenamic acid has the potential to increase the response of pancreatic adenocarcinoma to radiation therapy.

Tolfenamic acid inhibits esophageal cancer through repression of specificity proteins and c-Met.

The non-steroidal anti-inflammatory drug tolfenamic acid (TA) inhibits proliferation of SEG-1 and BIC-1 esophageal cancer cells with half-maximal growth inhibitory concentration values of 36 and 48 muM, respectively. TA also increased Annexin V staining in both cell lines, indicative of proapoptotic activity. Treatment of SEG-1 and BIC-1 cells with TA for up to 72 h decreased expression of specificity protein (Sp) transcription factors Sp1, Sp3 and Sp4 and this was accompanied by decreased expression of the well-characterized Sp-regulated genes cyclin D1, vascular endothelial growth factor and survivin. TA also decreased hepatocyte growth factor receptor, (c-Met), a receptor tyrosine kinase that is overexpressed in esophageal cancer cells and tumors and is an important drug target. Knockdown of Sp1, Sp3 and Sp4 by RNA interference in SEG-1 and BIC-1 cells also decreased c-Met expression, demonstrating that c-Met is an Sp-regulated gene in esophageal cancer cells. Sp1 was overexpressed in esophageal cancer cells and tumors and increased Sp1 staining was observed in esophageal tumors from patients. TA (20 mg/kg/day) also decreased tumor growth and weight in athymic nude mice bearing SEG-1 cells as xenografts and this was accompanied by increased apoptosis and decreased Sp1 and c-Met staining in tumors from treated mice. Thus, TA-dependent downregulation of Sp transcription factors and c-Met defines a novel chemotherapeutic approach for treatment of esophageal cancer.

Novel murine model for colon cancer: non-operative trans-anal rectal injection.

BACKGROUND: This study was conducted to develop a modified murine model of colon cancer that is non-operative. Currently, the most accurate orthotopic murine model of colon cancer requires an invasive procedure involving cecal injection of colon cancer cells and therefore limits the ability to perform immunological studies subsequent to cecal resections. MATERIALS AND METHODS: Murine colon cancer (CT26) cells were injected submucosally into the distal, posterior rectum of BALB/c mice. Care was taken not to pass transmurally into the pelvic cavity. Different magnifications (10x versus 100x) were used for injection, and primary tumor growth and metastatic disease were studied. RESULTS: In the initial study, 3/7 mice injected using 10x magnifications had notable, large tumor originating from the rectal wall, and histology revealed that all excised tumors were poorly differentiated adenocarcinoma. In the second study, 8/10 mice injected using 100x magnifications had notable tumor originating from the rectal well, and 4/8 mice had abnormal lung tissue with pathological evidence of hemorrhagic pulmonary edema. The use of 10x magnification resulted in 43% tumor take. In sharp contrast, 80% tumor take was observed with 100x magnification. The overall success of tumor take was 65% using the trans-anal rectal injection model. CONCLUSIONS: Our modified orthotopic murine model of colon cancer offers an alternative non-operative murine model for colon cancer and is less invasive than the traditional orthotopic model (i.e., cecal injection). This model may allow for more accurate investigations of inflammation and immune responses to surgical intervention without the influence of previous abdominal surgery.

Bcl-B expression in human epithelial and nonepithelial malignancies.

PURPOSE: Apoptosis plays an important role in neoplastic processes. Bcl-B is an antiapoptotic Bcl-2 family member, which is known to change its phenotype upon binding to Nur77/TR3. The expression pattern of this protein in human malignancies has not been reported. EXPERIMENTAL DESIGN: We investigated Bcl-B expression in normal human tissues and several types of human epithelial and nonepithelial malignancy by immunohistochemistry, correlating results with tumor stage, histologic grade, and patient survival. RESULTS: Bcl-B protein was strongly expressed in all normal plasma cells but found in only 18% of multiple myelomas (n = 133). Bcl-B immunostaining was also present in normal germinal center centroblasts and centrocytes and in approximately half of diffuse large B-cell lymphoma (n = 48) specimens, whereas follicular lymphomas (n = 57) did not contain Bcl-B. In breast (n = 119), prostate (n = 66), gastric (n = 180), and colorectal (n = 106) adenocarcinomas, as well as in non-small cell lung cancers (n = 82), tumor-specific overexpression of Bcl-B was observed. Bcl-B expression was associated with variables of poor prognosis, such as high tumor grade in breast cancer (P = 0.009), microsatellite stability (P = 0.0002), and left-sided anatomic location (P = 0.02) of colorectal cancers, as well as with greater incidence of death from prostate cancer (P = 0.005) and shorter survival of patients with small cell lung cancer (P = 0.009). Conversely, although overexpressed in many gastric cancers, Bcl-B tended to correlate with better outcome (P = 0.01) and more differentiated tumor histology (P < 0.0001). CONCLUSIONS: Tumor-specific alterations in Bcl-B expression may define subsets of nonepithelial and epithelial neoplasms with distinct clinical behaviors.

Protection from radiation-induced pneumonitis using cerium oxide nanoparticles.

In an effort to combat the harmful effects of radiation exposure, we propose that rare-earth cerium oxide (CeO(2)) nanoparticles (free-radical scavengers) protect normal tissue from radiation-induced damage. Preliminary studies suggest that these nanoparticles may be a therapeutic regenerative nanomedicine that will scavenge reactive oxygen species, which are responsible for radiation-induced cell damage. The effectiveness of CeO(2) nanoparticles in radiation protection in murine models during high-dose radiation exposure is investigated, with the ultimate goal of offering a new approach to radiation protection, using nanotechnology. We show that CeO(2) nanoparticles are well tolerated by live animals, and they prevent the onset of radiation-induced pneumonitis when delivered to live animals exposed to high doses of radiation. In the end, these studies provide a tremendous potential for radioprotection and can lead to significant benefits for the preservation of human health and the quality of life for humans receiving radiation therapy.

Regulation of vascular endothelial growth factor receptor-1 expression by specificity proteins 1, 3, and 4 in pancreatic cancer cells.

Vascular endothelial growth factor receptor-1 (VEGFR1) is expressed in cancer cell lines and tumors and, in pancreatic and colon cancer cells, activation of VEGFR1 is linked to increased tumor migration and invasiveness. Tolfenamic acid, a nonsteroidal anti-inflammatory drug, decreases Sp protein expression in Panc-1 and L3.6pl pancreatic cancer cells, and this was accompanied by decreased VEGFR1 protein and mRNA and decreased luciferase activity on cells transfected with constructs (pVEGFR1) containing VEGFR1 promoter inserts. Comparable results were obtained in pancreatic cancer cells transfected with small inhibitory RNAs for Sp1, Sp3, and Sp4 and all three proteins bound to GC-rich elements in the VEGFR1 promoter. These results show that VEGFR1 is regulated by Sp proteins and that treatment with tolfenamic acid decreases expression of this critical angiogenic factor. Moreover, in vitro studies in Panc-1 cells show that activation of VEGFR1 by VEGFB to increase mitogen-activated protein kinase 1/2 phosphorylation and cell migration on collagen-coated plates is also inhibited by tolfenamic acid. Thus, targeted degradation of Sp proteins is highly effective for inhibiting VEGFR1 and associated angiogenic responses in pancreatic cancer.

Successful inhibition of intracranial human glioblastoma multiforme xenograft growth via systemic adenoviral delivery of soluble endostatin and soluble vascular endothelial growth factor receptor-2: laboratory investigation.

OBJECT: Glioblastoma multiforme (GBM) is characterized by neovascularization, raising the question of whether angiogenic blockade may be a useful therapeutic strategy for this disease. It has been suggested, however, that, to be useful, angiogenic blockade must be persistent and at levels sufficient to overcome proangiogenic signals from tumor cells. In this report, the authors tested the hypothesis that sustained high concentrations of 2 different antiangiogenic proteins, delivered using a systemic gene therapy strategy, could inhibit the growth of established intracranial U87 human GBM xenografts in nude mice. METHODS: Mice harboring established U87 intracranial tumors received intravenous injections of adenoviral vectors encoding either the extracellular domain of vascular endothelial growth factor receptor-2-Fc fusion protein (Ad-VEGFR2-Fc) alone, soluble endostatin (Ad-ES) alone, a combination of Ad-VEGFR2-Fc and Ad-ES, or immunoglobulin 1-Fc (Ad-Fc) as a control. RESULTS: Three weeks after treatment, magnetic resonance imaging-based determination of tumor volume showed that treatment with Ad-VEGFR2-Fc, Ad-ES, or Ad-VEGFR2-Fc in combination with Ad-ES, produced 69, 59, and 74% growth inhibition, respectively. Bioluminescent monitoring of tumor growth revealed growth inhibition in the same treatment groups to be 62, 74, and 72%, respectively. Staining with proliferating cell nuclear antigen and with terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling showed reduced tumor cell proliferation and increased apoptosis in all antiangiogenic treatment groups. CONCLUSIONS: These results suggest that systemic delivery and sustained production of endostatin and soluble VEGFR2 can slow intracranial glial tumor growth by both reducing cell proliferation and increasing tumor apoptosis. This work adds further support to the concept of using antiangiogenesis therapy for intracranial GBM.