Department of Cancer Biology Faculty Papers Copyright (c) 2013 Thomas Jefferson University All rights reserved. http://jdc.jefferson.edu/cbfp Recent documents in Department of Cancer Biology Faculty Papers en-us Fri, 22 Feb 2013 16:37:36 PST 3600 Aleuria Aurantia Lectin (AAL)-Reactive Immunoglobulin G Rapidly Appears in Sera of Animals following Antigen Exposure. http://jdc.jefferson.edu/cbfp/32 http://jdc.jefferson.edu/cbfp/32 Mon, 19 Nov 2012 13:01:24 PST We have discovered an Aleuria Aurantia Lectin (AAL)-reactive immunoglobulin G (IgG) that naturally occurs in the circulation of rabbits and mice, following immune responses induced by various foreign antigens. AAL can specifically bind to fucose moieties on glycoproteins. However, most serum IgGs are poorly bound by AAL unless they are denatured or treated with glycosidase. In this study, using an immunogen-independent AAL-antibody microarray assay that we developed, we detected AAL-reactive IgG in the sera of all animals that had been immunized 1-2 weeks previously with various immunogens with and without adjuvants and developed immunogen-specific responses. All of these animals subsequently developed immunogen-specific immune responses. The kinetics of the production of AAL-reactive IgG in mice and rabbits were distinct from those of the immunogen-specific IgGs elicited in the same animals: they rose and fell within one to two weeks, and peaked between four to seven days after exposure, while immunogen-specific IgGs continued to rise during the same period. Mass spectrometric profiling of the Fc glycoforms of purified AAL-reactive IgGs indicates that these are mainly comprised of IgGs with core-fucosylated and either mono-or non-galactosylated Fc N-glycan structures. Our results suggest that AAL-reactive IgG could be a previously unrecognized IgG subset that is selectively produced at the onset of a humoral response.

]]> Songming Chen et al. Initial experience with Imacor hTEE-guided management of patients following transplant and mechanical circulatory support. http://jdc.jefferson.edu/cbfp/31 http://jdc.jefferson.edu/cbfp/31 Mon, 12 Nov 2012 08:26:37 PST We reviewed an initial experience using a miniaturized transesophageal echocardiography (TEE) probe (ImaCor, Garden City, NY) with 3 patients in whom Imacor hTEE intervention was used as a point of care device to manage extracorporeal membrane oxygenation cannula placement, cardiac hemodynamics, and post-op cardiac pathophysiology. The management of transplant or mechanical circulatory support (MCS) patients is especially challenging: transplanted hearts pose unique pathophysiological challenges, and MCS significantly alters pressure-volume-flow relationships. Real-time “hemodynamic” TEE (hTEE) can help provide effective management by direct visualization of cardiac filling and function and help guide hemodynamic management. In our initial experience, hTEE can provide point-of-care management following transplant or MCS.

]]> Christopher Kang et al. Regulation of lipogenesis by cyclin-dependent kinase 8-mediated control of SREBP-1. http://jdc.jefferson.edu/cbfp/30 http://jdc.jefferson.edu/cbfp/30 Fri, 03 Aug 2012 11:14:49 PDT Altered lipid metabolism underlies several major human diseases, including obesity and type 2 diabetes. However, lipid metabolism pathophysiology remains poorly understood at the molecular level. Insulin is the primary stimulator of hepatic lipogenesis through activation of the SREBP-1c transcription factor. Here we identified cyclin-dependent kinase 8 (CDK8) and its regulatory partner cyclin C (CycC) as negative regulators of the lipogenic pathway in Drosophila, mammalian hepatocytes, and mouse liver. The inhibitory effect of CDK8 and CycC on de novo lipogenesis was mediated through CDK8 phosphorylation of nuclear SREBP-1c at a conserved threonine residue. Phosphorylation by CDK8 enhanced SREBP-1c ubiquitination and protein degradation. Importantly, consistent with the physiologic regulation of lipid biosynthesis, CDK8 and CycC proteins were rapidly downregulated by feeding and insulin, resulting in decreased SREBP-1c phosphorylation. Moreover, overexpression of CycC efficiently suppressed insulin and feeding-induced lipogenic gene expression. Taken together, these results demonstrate that CDK8 and CycC function as evolutionarily conserved components of the insulin signaling pathway in regulating lipid homeostasis.

]]> Xiaoping Zhao et al. Genetic ablation of Cav1 differentially affects melanoma tumor growth and metastasis in mice: role of Cav1 in Shh heterotypic signaling and transendothelial migration. http://jdc.jefferson.edu/cbfp/29 http://jdc.jefferson.edu/cbfp/29 Fri, 27 Jul 2012 11:55:32 PDT Both cell-autonomous and non-cell-autonomous factors contribute to tumor growth and metastasis of melanoma. The function of caveolin-1 (Cav1), a multifunctional scaffold protein known to modulate several biologic processes in both normal tissue and cancer, has been recently investigated in melanoma cancer cells, but its role in the melanoma microenvironment remains largely unexplored. Here, we show that orthotopic implantation of B16F10 melanoma cells in the skin of Cav1KO mice increases tumor growth, and co-injection of Cav1-deficient dermal fibroblasts with melanoma cells is sufficient to recapitulate the tumor phenotype observed in Cav1KO mice. Using indirect coculture experiments with fibroblasts and melanoma cells combined with cytokine analysis, we found that Cav1-deficient fibroblasts promoted the growth of melanoma cells via enhanced paracrine cytokine signaling. Specifically, Cav1-deficient fibroblasts displayed increased ShhN expression, which heterotypically enhanced the Shh signaling pathway in melanoma cells. In contrast to primary tumor growth, the ability of B16F10 melanoma cells to form lung metastases was significantly reduced in Cav1KO mice. This phenotype was associated mechanistically with the inability of melanoma cells to adhere to and to transmigrate through a monolayer of endothelial cells lacking Cav1. Together, our findings show that Cav1 may regulate different mechanisms during primary melanoma tumor growth and metastatic dissemination.

]]> Franco Capozza et al. Signal transducer and activator of transcription-5 mediates neuronal apoptosis induced by inhibition of Rac GTPase activity. http://jdc.jefferson.edu/cbfp/28 http://jdc.jefferson.edu/cbfp/28 Thu, 05 Jul 2012 07:19:05 PDT In several neuronal cell types, the small GTPase Rac is essential for survival. We have shown previously that the Rho family GTPase inhibitor Clostridium difficile toxin B (ToxB) induces apoptosis in primary rat cerebellar granule neurons (CGNs) principally via inhibition of Rac GTPase function. In the present study, incubation with ToxB activated a proapoptotic Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway, and a pan-JAK inhibitor protected CGNs from Rac inhibition. STAT1 expression was induced by ToxB; however, CGNs from STAT1 knock-out mice succumbed to ToxB-induced apoptosis as readily as wild-type CGNs. STAT3 displayed enhanced tyrosine phosphorylation following treatment with ToxB, and a reputed inhibitor of STAT3, cucurbitacin (JSI-124), reduced CGN apoptosis. Unexpectedly, JSI-124 failed to block STAT3 phosphorylation, and CGNs were not protected from ToxB by other known STAT3 inhibitors. In contrast, STAT5A tyrosine phosphorylation induced by ToxB was suppressed by JSI-124. In addition, roscovitine similarly inhibited STAT5A phosphorylation and protected CGNs from ToxB-induced apoptosis. Consistent with these results, adenoviral infection with a dominant negative STAT5 mutant, but not wild-type STAT5, significantly decreased ToxB-induced apoptosis of CGNs. Finally, chromatin immunoprecipitation with a STAT5 antibody revealed increased STAT5 binding to the promoter region of prosurvival Bcl-xL. STAT5 was recruited to the Bcl-xL promoter region in a ToxB-dependent manner, and this DNA binding preceded Bcl-xL down-regulation, suggesting transcriptional repression. These data indicate that a novel JAK/STAT5 proapoptotic pathway significantly contributes to neuronal apoptosis induced by the inhibition of Rac GTPase.

]]> Trisha R Stankiewicz et al. The production of antibody by invading B cells is required for the clearance of rabies virus from the central nervous system. http://jdc.jefferson.edu/cbfp/27 http://jdc.jefferson.edu/cbfp/27 Thu, 08 Mar 2012 12:53:18 PST BACKGROUND: The pathogenesis of rabies is associated with the inability to deliver immune effectors across the blood-brain barrier and to clear virulent rabies virus from CNS tissues. However, the mechanisms that facilitate immune effector entry into CNS tissues are induced by infection with attenuated rabies virus.

METHODOLOGY/PRINCIPAL FINDINGS: Infection of normal mice with attenuated rabies virus but not immunization with killed virus can promote the clearance of pathogenic rabies virus from the CNS. T cell activity in B cell-deficient mice can control the replication of attenuated virus in the CNS, but viral mRNA persists. Low levels of passively administered rabies virus-neutralizing antibody reach infected cells in the cerebellum of B cell-deficient mice but are not sufficient to mediate virus clearance. Production of rabies virus-specific antibody by B cells invading CNS tissues is required for this process, and a substantial proportion of the B cells that accumulate in the CNS of mice infected with attenuated rabies virus produce virus-specific antibodies.

CONCLUSIONS/SIGNIFICANCE: The mechanisms required for immune effectors to enter rabies virus-infected tissues are induced by infection with attenuated rabies virus but not by infection with pathogenic rabies viruses or immunization with killed virus. T cell activities can inhibit rabies virus replication, but the production of rabies virus-specific antibodies by infiltrating B cells, as opposed to the leakage of circulating antibody across the BBB, is critical to elimination of the virus. These findings suggest that a pathogenic rabies virus infection may be treatable after the virus has reached the CNS tissues, providing that the appropriate immune effectors can be targeted to the infected tissues.

]]> D Craig Hooper et al. Animals Antibodies, Viral B-Lymphocytes Central Nervous System Disease Models, Animal Mice Mice, Inbred C57BL Rabies Rabies Vaccines Rabies virus Nerve Growth factor regulation of cyclin D1 in PC12 cells through a p21RAS extracellular signal-regulated kinase pathway requires cooperative interactions between Sp1 and nuclear factor-kappaB. http://jdc.jefferson.edu/cbfp/26 http://jdc.jefferson.edu/cbfp/26 Thu, 08 Mar 2012 12:53:17 PST The PC12 pheochromocytoma cell line responds to nerve growth factor (NGF) by exiting from the cell cycle and differentiating to induce extending neurites. Cyclin D1 is an important regulator of G1/S phase cell cycle progression, and it is known to play a role in myocyte differentiation in cultured cells. Herein, NGF induced cyclin D1 promoter, mRNA, and protein expression via the p21(RAS) pathway. Antisense- or small interfering RNA to cyclin D1 abolished NGF-mediated neurite outgrowth, demonstrating the essential role of cyclin D1 in NGF-mediated differentiation. Expression vectors encoding mutants of the Ras/mitogen-activated protein kinase pathway, and chemical inhibitors, demonstrated NGF induction of cyclin D1 involved cooperative interactions of extracellular signal-regulated kinase, p38, and phosphatidylinositol 3-kinase pathways downstream of p21(RAS). NGF induced the cyclin D1 promoter via Sp1, nuclear factor-kappaB, and cAMP-response element/activated transcription factor sites. NGF induction via Sp1 involved the formation of a Sp1/p50/p107 complex. Cyclin D1 induction by NGF governs differentiation and neurite outgrowth in PC12 cells.

]]> Francesco Marampon et al. Animals Base Sequence Cell Cycle Cell Proliferation Cyclin D1 Extracellular Signal-Regulated MAP Kinases Humans MAP Kinase Signaling System Mice NF-kappa B NF-kappa B p50 Subunit Nerve Growth Factor Neurites PC12 Cells Phosphatidylinositol 3-Kinases Promoter Regions, Genetic Protein Binding Proto-Oncogene Proteins p21(ras) RNA, Messenger RNA, Small Interfering Rats Retinoblastoma-Like Protein p107 Sp1 Transcription Factor Transcription, Genetic p38 Mitogen-Activated Protein Kinases Differential impact of tumor suppressor pathways on DNA damage response and therapy-induced transformation in a mouse primary cell model. http://jdc.jefferson.edu/cbfp/25 http://jdc.jefferson.edu/cbfp/25 Wed, 01 Feb 2012 12:47:39 PST The RB and p53 tumor suppressors are mediators of DNA damage response, and compound inactivation of RB and p53 is a common occurrence in human cancers. Surprisingly, their cooperation in DNA damage signaling in relation to tumorigenesis and therapeutic response remains enigmatic. In the context of individuals with heritable retinoblastoma, there is a predilection for secondary tumor development, which has been associated with the use of radiation-therapy to treat the primary tumor. Furthermore, while germline mutations of the p53 gene are critical drivers for cancer predisposition syndromes, it is postulated that extrinsic stresses play a major role in promoting varying tumor spectrums and disease severities. In light of these studies, we examined the tumor suppressor functions of these proteins when challenged by exposure to therapeutic stress. To examine the cooperation of RB and p53 in tumorigenesis, and in response to therapy-induced DNA damage, a combination of genetic deletion and dominant negative strategies was employed. Results indicate that loss/inactivation of RB and p53 is not sufficient for cellular transformation. However, these proteins played distinct roles in response to therapy-induced DNA damage and subsequent tumorigenesis. Specifically, RB status was critical for cellular response to damage and senescence, irrespective of p53 function. Loss of RB resulted in a dramatic evolution of gene expression as a result of alterations in epigenetic programming. Critically, the observed changes in gene expression have been specifically associated with tumorigenesis, and RB-deficient, recurred cells displayed oncogenic characteristics, as well as increased resistance to subsequent challenge with discrete therapeutic agents. Taken together, these findings indicate that tumor suppressor functions of RB and p53 are particularly manifest when challenged by cellular stress. In the face of such challenge, RB is a critical suppressor of tumorigenesis beyond p53, and RB-deficiency could promote significant cellular evolution, ultimately contributing to a more aggressive disease.

]]> A Kathleen McClendon et al. Animals Base Sequence Blotting, Western Cell Cycle Cell Transformation, Neoplastic Cells, Cultured DNA Damage DNA Primers Genes, Retinoblastoma Genes, Tumor Suppressor Genes, p53 Mice Models, Biological Mutation Oligonucleotide Array Sequence Analysis Reverse Transcriptase Polymerase Chain Reaction STAT5 regulation of BCL10 parallels constitutive NFkappaB activation in lymphoid tumor cells. http://jdc.jefferson.edu/cbfp/24 http://jdc.jefferson.edu/cbfp/24 Sun, 16 May 2010 18:06:03 PDT BACKGROUND: Signal Transducer and Activator of Transcription 5 A and B (STAT5) are key survival factors in cells of the lymphoid lineage. Identification of novel, tissue-specific STAT5 regulated genes would advance the ability to combat diseases due to aberrant STAT5 signaling. In the present work a library of human STAT5 bound genomic elements was created and validated. RESULTS: Of several STAT5 responsive genomic regulatory elements identified, one was located within the first intron of the human BCL10 gene. Chromatin immuno-precipitation reactions confirmed constitutive in vivo STAT5 binding to this intronic fragment in various human lymphoid tumor cell lines. Interestingly, non-phosphorylated STAT5 was found in the nuclei of Kit225 and YT cells in the absence of cytokine stimulation that paralleled constitutive NFkappaB activation. Inhibition of the hyperactive JAK3/STAT5 pathway in MT-2 cells via the Mannich-base, NC1153, diminished the constitutive in vivo occupancy of BCL10-SBR by STAT5, reduced NFkappaB activity and BCL10 protein expression in a dose dependent manner. Moreover, depletion of STAT5 via selective antisense oligonucleotide treatment similarly resulted in decreased BCL10 mRNA and protein expression, cellular viability and impaired NFkappaB activity independent of IL-2. CONCLUSION: These results suggest that the NFkappaB regulator BCL10 is an IL-2-independent STAT5 target gene. These findings proffer a model in which un-activated STAT5 can regulate pathways critical for lymphoid cell survival and inhibitors that disrupt STAT5 function independent of tyrosine phosphorylation may be therapeutically effective in treating certain leukemias/lymphomas.

]]> Zsuzsanna S Nagy et al. A novel and generalizable organotypic slice platform to evaluate stem cell potential for targeting pediatric brain tumors. http://jdc.jefferson.edu/cbfp/23 http://jdc.jefferson.edu/cbfp/23 Sun, 16 May 2010 17:35:47 PDT Brain tumors are now the leading cause of cancer-related deaths in children under age 15. Malignant gliomas are, for all practical purposes, incurable and new therapeutic approaches are desperately needed. One emerging strategy is to use the tumor tracking capacity inherent in many stem cell populations to deliver therapeutic agents to the brain cancer cells. Current limitations of the stem cell therapy strategy include that stem cells are treated as a single entity and lack of uniform technology is adopted for selection of clinically relevant sub-populations of stem cells. Specifically, therapeutic success relies on the selection of a clinically competent stem cell population based on their capacity of targeting brain tumors. A novel and generalizable organotypic slice platform to evaluate stem cell potential for targeting pediatric brain tumors is proposed to fill the gap in the current work flow of stem cell-based therapy. The organotypic slice platform has advantages of being mimic in vivo model, easier to manipulate to optimize parameters than in vivo models such as rodents and primates. This model serves as a framework to address the discrepancy between anticipated in vivo results and actual in vivo results, a critical barrier to timely progress in the field of the use of stem cells for the treatment of neurological disorders.

]]> Shengwen Calvin Li et al. High adsorption rate is detrimental to bacteriophage fitness in a biofilm-like environment. http://jdc.jefferson.edu/cbfp/22 http://jdc.jefferson.edu/cbfp/22 Thu, 01 Apr 2010 07:49:51 PDT BACKGROUND: Bacterial biofilm is ubiquitous in nature. However, it is not clear how this crowded habitat would impact the evolution of bacteriophage (phage) life history traits. In this study, we constructed isogenic lambda phage strains that only differed in their adsorption rates, because of the presence/absence of extra side tail fibers or improved tail fiber J, and maker states. The high cell density and viscosity of the biofilm environment was approximated by the standard double-layer agar plate. The phage infection cycle in the biofilm environment was decomposed into three stages: settlement on to the biofilm surface, production of phage progeny inside the biofilm, and emigration of phage progeny out of the current focus of infection.

RESULTS: We found that in all cases high adsorption rate is beneficial for phage settlement, but detrimental to phage production (in terms of plaque size and productivity) and emigration out of the current plaque. Overall, the advantage of high adsorption accrued during settlement is more than offset by the disadvantages experienced during the production and emigration stages. The advantage of low adsorption rate was further demonstrated by the rapid emergence of low-adsorption mutant from a high-adsorption phage strain with the side tail fibers. DNA sequencing showed that 19 out of the 21 independent mutant clones have mutations in the stf gene, with the majority of them being single-nucleotide insertion/deletion mutations occurring in regions with homonucleotide runs.

CONCLUSION: We conclude that high mutation rate of the stf gene would ensure the existence of side tail fiber polymorphism, thus contributing to rapid adaptation of the phage population between diametrically different habitats of benthic biofilm and planktonic liquid culture. Such adaptability would also help to explain the maintenance of the stf gene in phage lambda's genome.

]]> Romain Gallet et al. Stromal Caveolin-1 Levels Predict Early Ductal Carcinoma in Situ Progression to Invasive Breast Carcinoma http://jdc.jefferson.edu/cbfp/21 http://jdc.jefferson.edu/cbfp/21 Wed, 16 Dec 2009 10:32:13 PST With the increased use of screening mammography, the incidence of ductal carcinoma in situ (DCIS) has increased substantially and it currently accounts for 20-30% of newly diagnosed breast carcinomas in United States. Breast conserving therapy with or without radiotherapy is the accepted treatment for most cases of DCIS. However, the local recurrence rate with such therapy rages from 10 to 40% with half of these patients developing invasive carcinoma. Tamoxifen, used as an adjuvant systemic therapy for DCIS, does not increase overall survival and data on reduction of local recurrence rates are conflicting.

The current classification for DCIS based on nuclear grade, architectural differentiation and presence of necrosis, does not adequately predict the likelihood of recurrence after breast conserving therapy. Therefore, there is a critical need to identify novel predictors of DCIS progression and potential targets for therapy.

Recently, we showed that an absence of stromal (caveolin 1) Cav-1 expression in human breast cancers is a powerful single independent predictor of early disease recurrence, metastasis and poor clinical outcome. However, it remains unknown where stromal Cav-1 levels play any role in the progression of DCIS. The aim of this study was to evaluate the stromal expression of Cav-1 in a cohort of DCIS patients treated with wide-excision and close follow-up and examined the association between stromal Cav-1 expression and clinicopathological variables, DCIS recurrence or progression to invasive breast cancer.

]]> A. K. Witkiewicz et al. Plant Vaccines: An Immunological Perspective. http://jdc.jefferson.edu/cbfp/20 http://jdc.jefferson.edu/cbfp/20 Thu, 19 Nov 2009 06:34:08 PST The advent of technologies to express heterologous proteins in planta has led to the proposition that plants may be engineered to be safe, inexpensive vehicles for the production of vaccines and possibly even vectors for their delivery. The immunogenicity of a variety of antigens of relevance to vaccination expressed in different plants has been assessed. The purpose of this article is to examine the utility of plant-expression systems in vaccine development from an immunological perspective.

]]> Douglas C. Hooper Immune Evasion by Rabies Viruses through the Maintenance of Blood-Brain Barrier Integrity. http://jdc.jefferson.edu/cbfp/19 http://jdc.jefferson.edu/cbfp/19 Thu, 19 Nov 2009 06:26:49 PST The attenuated rabies virus (RV) strain Challenge Virus Standard (CVS)-F3 and a highly pathogenic strain associated with the silver-haired bats (SHBRV) can both be cleared from the central nervous system (CNS) tissues by appropriate antiviral immune mechanisms if the effectors are provided access across the blood-brain barrier (BBB). In the case of SHBRV infection, antiviral immunity develops normally in the periphery but fails to open the BBB, generally resulting in a lethal outcome. To determine whether or not an absence in the CNS targeted immune response is associated with the infection with other pathogenic RV strains, we have assessed the development of immunity, BBB permeability, and immune cell infiltration into the CNS tissues of mice infected with a variety of RV strains, including the dog variants responsible for the majority of human rabies cases. We demonstrate that the lethal outcomes of infection with a variety of known pathogenic RV strains are indeed associated with the inability to deliver immune effectors across the BBB. Survival from infection with certain of these viruses is improved in mice prone to CNS inflammation. The results suggest that competition between the activity of the immune effectors reaching CNS tissues and the inherent pathological attributes of the virus dictates the outcome and that intervention to deliver RV-specific immune effectors into CNS tissues may have general therapeutic value in rabies.

]]> Anirban Roy et al. Immunological assessment of plant-derived avian flu H5/HA1 variants. http://jdc.jefferson.edu/cbfp/18 http://jdc.jefferson.edu/cbfp/18 Mon, 23 Mar 2009 12:33:36 PDT Polypeptide variants of the HA1 antigenic domain of the H5N1 avian influenza virus hemagglutinin (HA) molecule were produced in plants using transient and stable expression systems and fused with His/c-myc tags or with mouse or human Fc antibody fragments. The resulting peptides were purified and used for intramuscular immunization of mice. While the recombinant HA1 variants induced a significant serum humoral immune response in the mice, none of the HA1 preparations induced virus-neutralizing antibodies. Fusion with the Fc fragment improved overall yield of the constructs and allowed purification requiring only a single step, but led to no detectable fusion-related enhancement of immunogenicity or quality of immune response.

]]> S Spitsin et al. Transcription factor signal transducer and activator of transcription 5 promotes growth of human prostate cancer cells in vivo http://jdc.jefferson.edu/cbfp/17 http://jdc.jefferson.edu/cbfp/17 Fri, 13 Mar 2009 12:49:50 PDT Purpose: Stat5a/b is the key mediator of prolactin (Prl) effects in prostate cancer cells via activation of Jak2. Prl is locally produced growth factor in human prostate cancer. Prl protein expression and constitutive activation of Stat5a/b are associated with high histological grade of clinical prostate cancer. Moreover, activation of Stat5a/b in primary prostate cancer predicts early disease recurrence. Here, we inhibited Stat5a/b by several different methodological approaches. Our goal was to establish a proof-of-principle that Stat5a/b is critical for prostate cancer cell viability in vitro and for prostate tumor growth in vivo.

Experimental Design: We inhibited Stat5a/b protein expression by antisense oligonucleotides or RNA interference and transcriptional activity of Stat5a/b by adenoviral expression of a dominant-negative mutant of Stat5a/b in prostate cancer cells in culture. Moreover, Stat5a/b activity was suppressed in human prostate cancer xenograft tumors in nude mice. Stat5a/b regulation of BclXL and Cyclin-D1 protein levels was demonstrated by antisense suppression of Stat5a/b protein expression followed by Western blotting.

Results and Conclusions: We show here that inhibition of Stat5a/b by antisense oligonuleotides, RNA interference, or adenoviral expression of DNStat5a/b all effectively kill prostate cancer cells. Moreover, we demonstrate that Stat5a/b is critical for human prostate cancer xenograft growth in nude mice. Stat5a/b effects on the viability of on prostate cancer cells involve Stat5a/b-regulation of BclXL and Cyclin-D1 protein levels, but not the expression or activation of Stat3. This work establishes Stat5a/b as a therapeutic target protein for prostate cancer. Pharmacological inhibition of Stat5a/b in prostate cancer can be achieved by small-molecule inhibitors of transactivation, dimerization or DNA-binding of Stat5a/b.

]]> Ayush Dagbadorj et al. Stabilization of SMAR1 mRNA by PGA2 involves a stem loop structure in the 5' UTR http://jdc.jefferson.edu/cbfp/16 http://jdc.jefferson.edu/cbfp/16 Mon, 29 Sep 2008 12:21:48 PDT Prostaglandins are anticancer agents known to inhibit tumor cell proliferation both in vitro and in vivo by affecting the mRNA stability. Here we report that a MAR-binding protein SMAR1 is a target of Prostaglandin A2 (PGA2) induced growth arrest. We identify a regulatory mechanism leading to stabilization of SMAR1 transcript. Our results show that a minor stem and loop structure present in the 5' UTR of SMAR1 (1-UTR) is critical for nucleoprotein complex formation that leads to SMAR1 stabilization in response to PGA2. This results in an increased SMAR1 transcript and altered protein levels, that in turn causes downregulation of Cyclin D1 gene, essential for G1/S phase transition. We also provide evidence for the presence of a variant 5' UTR SMAR1 (17-UTR) in breast cancer-derived cell lines. This form lacks the minor stem and loop structure required for mRNA stabilization in response to PGA2. As a consequence of this, there is a low level of endogenous tumor suppressor protein SMAR1 in breast cancer-derived cell lines. Our studies provide a mechanistic insight into the regulation of tumor suppressor protein SMAR1 by a cancer therapeutic PGA2, that leads to repression of Cyclin D1 gene.

]]> Lakshminarasimhan Pavritha et al. Plant-derived EpCAM antigen induces protective anti-cancer response. http://jdc.jefferson.edu/cbfp/15 http://jdc.jefferson.edu/cbfp/15 Tue, 10 Jun 2008 08:52:30 PDT Immunotherapy holds great promise for treatment of infectious and malignant diseases and might help to prevent the occurrence and recurrence of cancer. We produced a plant-derived tumor-associated colorectal cancer antigen EpCAM (pGA733) at high yields using two modern plant expression systems. The full antigenic domain of EpCAM was efficiently purified to confirm its antigenic and immunogenic properties as compared to those of the antigen expressed in the baculovirus system (bGA733). Recombinant plant-derived antigen induced a humoral immune response in BALB/c mice. Sera from those mice efficiently inhibited the growth of SW948 colorectal carcinoma cells xenografted in nude mice, as compared to the EpCAM-specific mAb CO17-1A. Our results support the feasibility of producing anti-cancer recombinant vaccines using plant expression systems.

]]> Robert Brodzik et al. Micro RNA 145 targets the insulin receptor substrate-1 and inhibits the growth of colon cancer cells http://jdc.jefferson.edu/cbfp/14 http://jdc.jefferson.edu/cbfp/14 Fri, 11 Apr 2008 13:53:08 PDT The insulin receptor substrate-1 (IRS-1), a docking protein for both the type 1 insulin-like growth factor receptor (IGF-IR) and the insulin receptor, is known to send a mitogenic, anti-apoptotic, and anti-differentiation signal. Several micro RNAs (miRs) are suggested by the data base as possible candidates for targeting IRS-1. We show here that one of the miRs predicted by the data base, miR145, whether transfected as a synthetic oligonucleotide or expressed from a plasmid, causes down-regulation of IRS-1 in human colon cancer cells. IRS-1 mRNA is not decreased by miR145, while it is down-regulated by an siRNA targeting IRS-1. Targeting of the IRS-1 3'-untranslated region (UTR) by miR145 was confirmed using a reporter gene (luciferase) expressing the miR145 binding sites of the IRS-1 3'-UTR. In agreement with the role of IRS-1 in cell proliferation, we show that treatment of human colon cancer cells with miR145 causes growth arrest comparable to the use of an siRNA against IRS-1. Taken together, these results identify miR145 as a micro RNA that down-regulates the IRS-1 protein, and inhibits the growth of human cancer cells.

]]> Bin Shi et al. Sirtuins, nuclear hormone receptor acetylation and transcriptional regulation http://jdc.jefferson.edu/cbfp/13 http://jdc.jefferson.edu/cbfp/13 Wed, 05 Mar 2008 11:09:24 PST Endocrine signaling via nuclear receptors (NRs) is known to play an important role in normal physiology as well as in human tumor progression. Hormones regulate gene expression by altering local chromatin structure and, thereby, accessibility of transcriptional co-regulators to DNA. Recently it has been shown that non-histone proteins involved in hormone signaling, such as nuclear receptors and NR co-activators, are regulated by acetylation, resulting in their altered transcriptional activity. NAD-dependent protein deacetylases, the sirtuins (Sir2-related enzymes), directly modify NRs. Because sirtuins have been shown to regulate tumor cellular growth, aging, metabolic signaling and endocrine hormone signaling, they might play a role in cancer progression. This review focuses on the role of acetylation and the sirtuins in nuclear hormone receptor signaling.

]]> James R. Whittle et al.