RESUMEN
Overexpressed tumor-associated antigens (TAAs) are a large group that includes proteins found at increased levels in tumors compared to healthy cells. Universal tumor expression can be defined as overexpression in all cancers examined as has been shown for Tumor Protein D52. TPD52 is an over expressed TAA actively involved in transformation, leading to increased proliferation and metastasis. TPD52 overexpression has been demonstrated in many human adult and pediatric malignancies. The murine orthologue of TPD52 (mD52) parallels normal tissue expression patterns and known functions of human TPD52 (hD52). Here in we present our preclinical studies over the past 15 years which have demonstrated that vaccine induced immunity against mD52 is effective against multiple cancers in murine models, without inducing autoimmunity against healthy tissues and cells.
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Vacunas contra el Cáncer , Neoplasias , Adulto , Niño , Humanos , Animales , Ratones , Proteínas de Neoplasias , Neoplasias/prevención & control , Antígenos de Neoplasias , Autoinmunidad , Línea Celular TumoralRESUMEN
Transplantation is a clinical procedure that treats a variety of diseases yet is unattainable for many patients due to a nationwide organ shortage and the harsh side effects of chronic immune suppression. Xenografted pig organs are an attractive alternative to traditional allografts and would provide an endless supply of transplantable tissue, but transplants risk rejection by the recipient's immune system. An essential component of the rejection immune response is the complement system. Sertoli cells, an immunoregulatory testicular cell, survive complement as xenografts long term without any immune suppressants. We hypothesized that exposure to the xenogeneic complement influences Sertoli cell gene expression of other accommodation factors that contribute to their survival; thus, the purpose of this study was to describe these potential changes in gene expression. RNA sequencing of baseline neonatal pig Sertoli cells (NPSC) as compared to NPSC after exposure to normal human serum (NHS, containing complement) revealed 62 significantly differentially expressed genes (DEG) that affect over 30 pathways involved in immune regulation, cell survival, and transplant accommodation. Twelve genes of interest were selected for further study, and Sertoli cell protein expression of CCL2 and the accommodation factor A20 were confirmed for the first time. Functional pathway analyses were conducted in NPSC and three biological clusters were revealed as being considerably affected by NHS exposure: innate immune signaling, cytokine signaling, and T cell regulation. Better understanding of the interaction of Sertoli cells with complement in a xenograft environment may reveal the mechanisms behind immune-privileged systems to increase graft viability.
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Cancer immunotherapy is a powerful tool for inducing antigen-specific antitumor cytotoxic T lymphocytes (CTLs). Next-generation strategies may include vaccination against overexpressed oncogenic tumor-self antigens. Previously, we reported vaccination against the oncogenic tumor-self antigen D52 (D52) was effective in preventing tumor growth. We recently reported that D52-vaccinated IL-10-deficient mice generated a significant memory response against tumor recurrence compared to wild-type mice and that vaccine-induced CD8+ IL-10+ T cells may possess regulatory function. Herein, we extended these studies by testing the hypothesis that D52-vaccine-elicited CD8+ IL-10+ T cells represent a distinct T cell population with a regulatory phenotype. C57Black/6J mice deficient in IL-10 or IFN-γ were vaccinated with the murine orthologue of D52; vaccination of wild-type (wt) mice served as a control for comparison. T cells were isolated from all three groups of vaccinated mice, and RNA was extracted from purified CD8+ T cells for deep sequencing and expression analysis. Chemokine receptor 8 (CCR8) and inducible co-stimulator (ICOS) were overexpressed in CD8+ T cells that produced IL-10 but not IFN-γ. These surface markers are associated with IL-10 producing CD4+ T regulatory cells thus supporting the possibility that CD8+ IL-10+ T cells elicited by D52 vaccination represent a unique regulatory T cell subset. The current phenotypic analyses of D52 vaccine elicited CD8+ T cells strengthen our premise that CD8+ IL-10+ T cells elicited by D52 tumor-self protein vaccination likely contribute to the suppression of memory CTL responses and inhibition of durable tumor immunity.
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Vacunas contra el Cáncer , Neoplasias , Ratones , Animales , Interleucina-10 , Autoantígenos/metabolismo , Linfocitos T CD8-positivos , Linfocitos T CD4-Positivos , Antígenos de Neoplasias/genética , Vacunación , Neoplasias/metabolismoRESUMEN
Dishevelled (DVL) critically regulates Wnt signaling and contributes to a wide spectrum of diseases and is important in normal and pathophysiological settings. However, how it mediates diverse cellular functions remains poorly understood. Recent discoveries have revealed that constitutive Wnt pathway activation contributes to breast cancer malignancy, but the mechanisms by which this occurs are unknown and very few studies have examined the nuclear role of DVL. Here, we have performed DVL3 ChIP-seq analyses and identify novel target genes bound by DVL3. We show that DVL3 depletion alters KMT2D binding to novel targets and changes their epigenetic marks and mRNA levels. We further demonstrate that DVL3 inhibition leads to decreased tumor growth in two different breast cancer models in vivo. Our data uncover new DVL3 functions through its regulation of multiple genes involved in developmental biology, antigen presentation, metabolism, chromatin remodeling, and tumorigenesis. Overall, our study provides unique insight into the function of nuclear DVL, which helps to define its role in mediating aberrant Wnt signaling.
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Neoplasias , Vía de Señalización Wnt , Proteínas Dishevelled/genética , Proteínas Dishevelled/metabolismo , Humanos , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Secuencias Reguladoras de Ácidos Nucleicos , Vía de Señalización Wnt/genéticaRESUMEN
Development of cancer vaccines targeting tumor self-antigens is complex and challenging due to the difficulty of overcoming immune tolerance to self-proteins. Vaccination against tumor self-protein D52 (D52) has been successful, although complete protection appears impaired by immune regulation. Our previous studies suggest that vaccine elicited CD8 + T cells producing interleukin 10 (IL-10) may have a negative impact on tumor protection. Understanding the role CD8+ IL-10 + T cells play in the immune response following vaccination with D52 could result in a more potent vaccine. To address this, we vaccinated IL-10 deficient mice with the murine orthologue of D52; vaccination of wild type (wt) C57BL/6J served as a control for comparison. In separate experiments, D52 vaccinated wt mice were administered IL-10R-specific mAb to neutralize IL-10 function. Interestingly, we observed similar protection against primary tumor challenge in the experimental groups compared to the controls. However, individual IL-10 deficient mice that rejected the primary tumor challenge were re-challenged 140 days post-primary challenge to access vaccine durability and immunologic memory against tumor recurrence. Mice deficient in IL-10 demonstrated a memory response in which 100% of the mice were protected from secondary tumor challenge, while wt mice had diminished recall response (25%) against tumor recurrence. These results with analysis of vaccine-elicited CD8 + T cells for tumor-specific killing and regulatory cell marker expression, add further support to our premise that CD8+ IL-10 + T cells elicited by D52 tumor-self protein vaccine contribute to the suppression of a memory CTL responses and durable tumor immunity.
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Linfocitos T CD8-positivos , Vacunas contra el Cáncer/inmunología , Proteínas de Neoplasias/inmunología , Recurrencia Local de Neoplasia , Animales , Interleucina-10 , Ratones , Ratones Endogámicos C57BL , VacunaciónRESUMEN
Tumor protein D52 (TPD52) is amplified and overexpressed in breast and prostate cancers which are frequently characterised by dysregulated lipid storage and metabolism. TPD52 expression increases lipid storage in mouse 3T3 fibroblasts, and co-distributes with the Golgi marker GM130 and lipid droplets (LDs). We examined the effects of Brefeldin A (BFA), a fungal metabolite known to disrupt the Golgi structure, in TPD52-expressing 3T3 cells, and in human AU565 and HMC-1-8 breast cancer cells that endogenously express TPD52. Five-hour BFA treatment reduced median LD numbers, but increased LD sizes. TPD52 knockdown decreased both LD sizes and numbers, and blunted BFA's effects on LD numbers. Following BFA treatment for 1-3 hours, TPD52 co-localised with the trans-Golgi network protein syntaxin 6, but after 5 hours BFA treatment, TPD52 showed increased co-localisation with LDs, which was disrupted by microtubule depolymerising agent nocodazole. BFA treatment also increased perilipin (PLIN) family protein PLIN3 but reduced PLIN2 detection at LDs in TPD52-expressing 3T3 cells, with PLIN3 recruitment to LDs preceding that of TPD52. An N-terminally deleted HA-TPD52 mutant (residues 40-184) almost exclusively targeted to LDs in both vehicle and BFA treated cells. In summary, delayed recruitment of TPD52 to LDs suggests that TPD52 participates in a temporal hierarchy of LD-associated proteins that responds to altered LD packaging requirements induced by BFA treatment.
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Brefeldino A/farmacología , Proteínas Asociadas a Gotas Lipídicas/metabolismo , Gotas Lipídicas/metabolismo , Metabolismo de los Lípidos , Proteínas de Neoplasias/metabolismo , Secuencia de Aminoácidos , Animales , Técnica del Anticuerpo Fluorescente , Técnicas de Silenciamiento del Gen , Aparato de Golgi/metabolismo , Ratones , Mutación , Proteínas de Neoplasias/genética , Perilipina-3/metabolismo , Transporte de ProteínasRESUMEN
Self-floating hollow glass microspheres (HGMS) modified with tumor-specific antibodies have been developed for the capture of circulating tumor cells (CTCs), and have demonstrated effective cell isolation and good viability of isolated cancer cells. However, the capture efficiency decreases dramatically if the spiked cell concentration is low, possibly due to insufficient interactions between cancer cells and the HGMS surface. In order to apply HGMS-based CTC isolation to clinically relevant samples, it is desirable to create nanostructures on the surface of HGMS to enhance cell-surface interactions. Nevertheless, current microfabrication methods cannot generate nanostructured-surfaces on microspheres. The authors have developed a new HGMS with a controlled nanotopographical surface structure (NSHGMS), and demonstrated isolation and recovery of rare cancer cells. NSHGMS are achieved by applying layer-by-layer (LbL) assembly of negatively charged SiO2 nanoparticles and positively charged poly-l-arginine molecules, then sheathing the surface with an enzymatically degradable LbL film made from biotinylated alginate and poly-l-arginine, and capping with anti-EpCAM antibodies and anti-fouling PEG molecules. Compared to smooth-surfaced HGMS, NSHGMS showed shorter isolation time (20 min), enhanced capture efficiency (93.6 ± 4.9%) and lower detection limit (30 cells per mL) for commonly used cancer cell lines (MCF7, SK-BR-3, PC-3, A549 and CCRF-CEM). This NSHGMS-based CTC isolation method does not require specialized lab equipment or an external power source, and thus, can be used for the separation of targeted cells from blood or other body fluids in a resource-limited environment.
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Separación Celular/métodos , Microesferas , Células Neoplásicas Circulantes , Línea Celular Tumoral , Humanos , Dióxido de SilicioRESUMEN
Tumor protein D52 (TPD52) is amplified and/or overexpressed in cancers of diverse cellular origins. Altered cellular metabolism (including lipogenesis) is a hallmark of cancer development, and protein-protein associations between TPD52 and known regulators of lipid storage, and differential TPD52 expression in obese versus non-obese adipose tissue, suggest that TPD52 might regulate cellular lipid metabolism. We found increased lipid droplet numbers in BALB/c 3T3 cell lines stably expressing TPD52, compared with control and TPD52L1-expressing cell lines. TPD52-expressing 3T3 cells showed increased fatty acid storage in triglyceride (from both de novo synthesis and uptake) and formed greater numbers of lipid droplets upon oleic acid supplementation than control cells. TPD52 colocalised with Golgi, but not endoplasmic reticulum (ER), markers and also showed partial colocalisation with lipid droplets coated with ADRP (also known as PLIN2), with a proportion of TPD52 being detected in the lipid droplet fraction. Direct interactions between ADRP and TPD52, but not TPD52L1, were demonstrated using the yeast two-hybrid system, with ADRP-TPD52 interactions confirmed using GST pulldown assays. Our findings uncover a new isoform-specific role for TPD52 in promoting intracellular lipid storage, which might be relevant to TPD52 overexpression in cancer.
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Ácidos Grasos/metabolismo , Regulación de la Expresión Génica/fisiología , Proteínas de Neoplasias/biosíntesis , Triglicéridos/metabolismo , Animales , Células 3T3 BALB , Línea Celular Tumoral , Retículo Endoplásmico/genética , Retículo Endoplásmico/metabolismo , Ácidos Grasos/genética , Femenino , Aparato de Golgi/genética , Aparato de Golgi/metabolismo , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Proteínas de Neoplasias/genética , Perilipina-2 , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Triglicéridos/genéticaRESUMEN
Overexpressed tumor-self antigens represent the largest group of candidate vaccine targets. Those exhibiting a role in oncogenesis may be some of the least studied but perhaps most promising. This review considers this subset of self antigens by highlighting vaccine efforts for some of the better known members and focusing on TPD52, a new promising vaccine target. We shed light on the importance of both preclinical and clinical vaccine studies demonstrating that tolerance and autoimmunity (presumed to preclude this class of antigens from vaccine development) can be overcome and do not present the obstacle that might have been expected. The potential of this class of antigens for broad application is considered, possibly in the context of low tumor burden or adjuvant therapy, as is the need to understand mechanisms of tolerance that are relatively understudied.
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Antígenos de Neoplasias/inmunología , Autoantígenos/inmunología , Vacunas contra el Cáncer/inmunología , Neoplasias/inmunología , Antígenos de Neoplasias/biosíntesis , Autoinmunidad/inmunología , Humanos , Tolerancia Inmunológica/inmunología , Inmunoterapia , Proteínas de Neoplasias/inmunología , Neoplasias/prevención & control , Neoplasias/terapia , Linfocitos T Citotóxicos/inmunología , Escape del Tumor/inmunologíaRESUMEN
The Tumor protein D52 (TPD52) gene was identified nearly 20 years ago through its overexpression in human cancer, and a substantial body of data now strongly supports TPD52 representing a gene amplification target at chromosome 8q21.13. This review updates progress toward understanding the significance of TPD52 overexpression and targeting, both in tumors known to be characterized by TPD52 overexpression/amplification, and those where TPD52 overexpression/amplification has been recently or variably reported. We highlight recent findings supporting microRNA regulation of TPD52 expression in experimental systems and describe progress toward deciphering TPD52's cellular functions, particularly in cancer cells. Finally, we provide an overview of TPD52's potential as a cancer biomarker and immunotherapeutic target. These combined studies highlight the potential value of genes such as TPD52, which are overexpressed in many cancer types, but have been relatively understudied.
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Proteínas de Neoplasias/fisiología , Oncogenes , Animales , Biomarcadores de Tumor/sangre , Línea Celular Tumoral , Supervivencia Celular , Cromosomas Humanos Par 8 , Daño del ADN , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica , Humanos , MicroARNs/fisiología , Mutación , Proteínas de Neoplasias/sangre , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/inmunología , VacunaciónRESUMEN
TPD52 and ERBB2 co-expression has been persistently reported in human breast cancer and animal models of this disease, but the significance of this is unknown. We identified significant positive associations between relative TPD52 and ERBB2 transcript levels in human diagnostic breast cancer samples, and maximal TPD52 expression in the hormone receptor (HR)- and ERBB2-positive sub-group. High-level TPD52 expression was associated with significantly reduced metastasis-free survival, within the overall cohort (log rank test, P = 8.6 × 10(-4), n = 375) where this was an independent predictor of metastasis-free survival (hazard ratio, 2.69, 95% confidence interval 1.59-4.54, P = 2.2 × 10(-4), n = 359), and the HR- and ERBB2-positive sub-group (log rank test, P = 0.035, n = 47). Transient TPD52 knock-down in the ERBB2-amplified breast cancer cell lines SK-BR-3 and BT-474 produced significant apoptosis, both singly and in combination with transient ERBB2 knock-down. Unlike ERBB2 knock-down, transient TPD52 knock-down produced no reduction in pAKT levels in SK-BR-3 or BT-474 cells. We then derived multiple SK-BR-3 cell lines in which TPD52 levels were stably reduced, and measured significant inverse correlations between pERBB2 and TPD52 levels in viable TPD52-depleted and control cell lines, all of which showed similar proliferative capacities. Our results therefore identify TPD52 as a survival factor in ERBB2-amplified breast cancer cells, and suggest complementary cellular functions for TPD52 and ERBB2.
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Neoplasias de la Mama/metabolismo , Proteínas de Neoplasias/fisiología , Receptor ErbB-2/genética , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias de la Mama/mortalidad , Neoplasias de la Mama/patología , Línea Celular Tumoral , Proliferación Celular , Supervivencia sin Enfermedad , Femenino , Amplificación de Genes , Expresión Génica , Humanos , Estimación de Kaplan-Meier , Metástasis Linfática , Persona de Mediana Edad , Modelos de Riesgos Proporcionales , Transporte de ProteínasRESUMEN
Tumor protein D52 (D52) is constitutively expressed in healthy tissues and overexpressed in multiple cancers, including (but not limited to) breast, prostate and ovarian carcinomas. Although the normal functions of D52 are unknown, it is clear that increased D52 expression levels not only stimulate cell proliferation and metastasis, but also correlate with poor prognosis in a subset of breast cancer patients. The murine orthologs of D52 (mD52) shares 86% identity with its human counterpart (hD52) and mirrors hD52 expression patterns. The forced overexpression of mD52 induces anchorage-independent growth in vitro and promotes tumor formation as well as spontaneous metastasis in vivo. We have previously reported that the intramuscular administration of recombinant mD52 elicits immune responses capable of rejecting a challenge with tumor cells and preventing spontaneous metastasis only in 50% of mice. We hypothesized that mechanisms of peripheral tolerance dampen immune responses against mD52, thus limiting the protective effects of vaccination. To test this hypothesis, mice were depleted of CD25+ regulatory T cells (Tregs) and subcutaneously immunized with mD52 prior to a tumor challenge. The subcutaneous immunization failed to induce protective antitumor immunity unless accompanied by Treg depletion, which resulted in a rate of protection of 70% as compared with.
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The central role of CD4+ T lymphocytes in mediating DNA vaccine-induced tumor immunity against the viral oncoprotein simian virus 40 (SV40) large tumor antigen (Tag) has previously been described by our laboratory. In the present study, we extend our previous findings by examining the roles of IFN-γ and Th1-associated effector cells within the context of DNA immunization in a murine model of pulmonary metastasis. Immunization of BALB/c mice with plasmid DNA encoding SV40 Tag (pCMV-Tag) generated IFN-γ-secreting T lymphocytes that produced this cytokine upon in vitro stimulation with mKSA tumor cells. The role of IFN-γ as a mediator of protection against mKSA tumor development was assessed via in vivo IFN-γ neutralization, and these experiments demonstrated a requirement for this cytokine in the induction immune phase. Neutralization of IFN-γ was associated with a reduction in Th1 cytokine-producing CD4+ and CD8+ splenocytes, as assessed by flow cytometry analysis, and provided further evidence for the role of CD4+ T lymphocytes as drivers of the cellular immune response. Depletion of NK cells and CD8+ T lymphocytes demonstrated the expendability of these cell types individually, but showed a requirement for a resident cytotoxic cell population within the immune effector phase. Our findings demonstrate the importance of IFN-γ in the induction of protective immunity stimulated by pCMV-Tag DNA-based vaccine and help to clarify the general mechanisms by which DNA vaccines trigger immunity to tumor cells.
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Antígenos Transformadores de Poliomavirus/inmunología , Interferón gamma/inmunología , Infecciones por Polyomavirus/inmunología , Virus 40 de los Simios/inmunología , Infecciones Tumorales por Virus/inmunología , Vacunas de ADN/inmunología , Animales , Antígenos Transformadores de Poliomavirus/genética , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/inmunología , Células Cultivadas , Femenino , Interferón gamma/metabolismo , Neoplasias Renales/inmunología , Neoplasias Renales/patología , Células Asesinas Naturales/inmunología , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/secundario , Ratones , Ratones Endogámicos BALB C , Plásmidos , Infecciones por Polyomavirus/genética , Bazo/inmunología , Células TH1/inmunología , Infecciones Tumorales por Virus/genéticaRESUMEN
A mechanistic analysis of tumor immunity directed toward the viral oncoprotein simian virus 40 (SV40) large tumor antigen (Tag) has previously been described by our laboratory for scenarios of recombinant Tag immunization in BALB/c mice. In the present study, we performed a preliminary characterization of the immune components necessary for systemic tumor immunity induced upon immunization with plasmid DNA encoding SV40 Tag as a transgene (pCMV-Tag). Antibody responses to SV40 Tag were observed via indirect enzyme-linked immunosorbent assay following three intramuscular (i.m.) injections of pCMV-Tag and were typified by a mixed Th1/Th2 response. Complete tumor immunity within a murine model of pulmonary metastasis was achieved upon two i.m. injections of pCMV-Tag, as assessed by examination of tumor foci in mouse lungs, without a detectable antibody response to SV40 Tag. Induction-phase and effector-phase depletions of T cell subsets were performed in vivo via administration of depleting rat monoclonal antibodies, and these experiments demonstrated that CD4(+) T lymphocytes are required in both phases of the adaptive immune response. Conversely, depletion of CD8(+) T lymphocytes did not impair tumor immunity in either immune phase and resulted in the premature production of antibodies to SV40 Tag. Our findings are unique in that a dominant role could be ascribed to CD4(+) T lymphocytes within a model of DNA vaccine-induced tumor immunity to Tag-expressing tumor cells. Additionally, our findings provide insight into the general mechanisms of vaccine-induced tumor immunity directed toward tumors bearing distinct tumor-associated antigens.
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Antígenos Virales de Tumores/inmunología , Linfocitos T CD4-Positivos/inmunología , Neoplasias Experimentales/inmunología , Plásmidos , Virus 40 de los Simios/inmunología , Vacunas de ADN/inmunología , Animales , Anticuerpos Antivirales/biosíntesis , Ensayo de Inmunoadsorción Enzimática , Femenino , Ratones , Ratones Endogámicos BALB C , Vacunas de ADN/administración & dosificaciónRESUMEN
The required activities of CD4(+) T cells and antibody against the virally encoded oncoprotein simian virus 40 (SV40) Tag have previously been demonstrated by our laboratory to be mediators in achieving antitumor responses and tumor protection through antibody-dependent cell-mediated cytotoxicity (ADCC). In this study, we further characterize the necessary immune cell components that lead to systemic tumor immunity within an experimental pulmonary metastatic model as the result of SV40 Tag immunization and antibody production. Immunized animals depleted of CD8(+) T cells at the onset of experimental tumor cell challenge developed lung tumor foci and had an overall decreased survival due to lung tumor burden, suggesting a role for CD8(+) T cells in the effector phase of the immune response. Lymphocytes and splenocytes harvested from SV40 Tag-immunized mice experimentally inoculated with tumor cells synthesized increased in vitro levels of the Th1 cytokine gamma interferon (IFN-gamma), as assessed by enzyme-linked immunosorbent assay (ELISA) and flow cytometry assays. CD8(+) T-cell activity was also heightened in SV40 Tag-immunized and tumor cell-challenged mice, based upon intracellular production of perforin, confirming the cytolytic properties of CD8(+) T cells against tumor cell challenge. Altogether, these data point to the role of recombinant SV40 Tag protein immunization in initiating a cytotoxic T-lymphocyte (CTL) response during tumor cell dissemination and growth. The downstream activity of CD8(+) T cells within this model is likely initiated from SV40 Tag-specific antibody mediating ADCC tumor cell destruction.
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Antígenos Transformadores de Poliomavirus/inmunología , Linfocitos T CD8-positivos/inmunología , Neoplasias Pulmonares/inmunología , Virus 40 de los Simios/inmunología , Infecciones Tumorales por Virus/inmunología , Animales , Anticuerpos Antivirales/sangre , Antígenos Transformadores de Poliomavirus/administración & dosificación , Línea Celular Transformada , Inmunidad Humoral , Inmunización , Riñón/citología , Riñón/virología , Neoplasias Pulmonares/mortalidad , Neoplasias Pulmonares/prevención & control , Depleción Linfocítica , Ratones , Ratones Endogámicos BALB C , Trasplante de Neoplasias , Linfocitos T Citotóxicos , Células TH1/inmunología , Infecciones Tumorales por Virus/mortalidad , Infecciones Tumorales por Virus/prevención & controlRESUMEN
Tumor protein D52 (TPD52) is involved in cellular transformation, proliferation and metastasis. TPD52 over expression has been demonstrated in several cancers including prostate, breast, and ovarian carcinomas. Murine TPD52 (mD52) has been shown to induce anchorage independent growth in vitro and metastasis in vivo, and mirrors the function and normal tissue expression patterns of the human orthologue of TPD52. We believe TPD52 represents a self, non-mutated tumor associated antigen (TAA) important for maintaining a transformed and metastatic cellular phenotype. The transgenic adeno-carcinoma of the mouse prostate (TRAMP) model was employed to study mD52 as a vaccine antigen. Naïve mice were immunized with either recombinant mD52 protein or plasmid DNA encoding the full-length cDNA of mD52. Following immunization, mice were challenged with a subcutaneous, tumorigenic dose of mD52 positive, autochthonous TRAMP-C1 tumor cells. Sixty percent of mice were tumor free 85 days post challenge with TRAMP-C1 when immunized with mD52 as a DNA-based vaccine admixed with soluble granulocyte-macrophage colony stimulating factor (GM-CSF). Survivors of the initial tumor challenge rejected a second tumor challenge given in the opposite flank approximately 150 days after the first challenge, and remained tumor free for more than an additional 100 days. The T cell cytokine secretion patterns from tumor challenge survivors indicated that a T(H)1-type cellular immune response was involved in tumor protection. These data suggest that mD52 vaccination induced a memory, cellular immune response that resulted in protection from murine prostate tumors that naturally over express mD52 protein.
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Adenocarcinoma/terapia , Autoantígenos/inmunología , Vacunas contra el Cáncer/uso terapéutico , Factor Estimulante de Colonias de Granulocitos y Macrófagos/administración & dosificación , Proteínas de Neoplasias/inmunología , Neoplasias de la Próstata/inmunología , Neoplasias de la Próstata/terapia , Vacunas de ADN/uso terapéutico , Adenocarcinoma/inmunología , Adenocarcinoma/prevención & control , Animales , Autoantígenos/genética , Vacunas contra el Cáncer/genética , Vacunas contra el Cáncer/inmunología , Línea Celular Tumoral , Citocinas/inmunología , Citocinas/metabolismo , Memoria Inmunológica/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas de Neoplasias/genética , Neoplasias de la Próstata/prevención & control , Proteínas Recombinantes , Linfocitos T Citotóxicos/inmunología , Células TH1/inmunología , Células TH1/metabolismo , Vacunación , Vacunas de ADN/genética , Vacunas de ADN/inmunologíaRESUMEN
PURPOSE: Tumor protein D52 (TPD52 or D52) is frequently overexpressed in breast and other cancers and present at increased gene copy number. It is, however, unclear whether D52 amplification and overexpression target specific functional properties of the encoded protein. EXPERIMENTAL DESIGN: The expression of D52-like genes and MAL2 was compared in breast tissues using quantitative reverse transcription-PCR. The functions of human D52 and D53 genes were then compared by stable expression in BALB/c 3T3 fibroblasts and transient gene knockdown in breast carcinoma cell lines. In situ D52 and MAL2 protein expression was analyzed in breast tissue samples using tissue microarray sections. RESULTS: The D52 (8q21.13), D54 (20q13.33), and MAL2 (8q24.12) genes were significantly overexpressed in breast cancer tissue (n = 95) relative to normal breast (n = 7; P = 0.005) unlike the D53 gene (6q22.31; P = 0.884). Subsequently, D52-expressing but not D53-expressing 3T3 cell lines showed increased proliferation and anchorage-independent growth capacity, and reduced D52 but not D53 expression in SK-BR-3 cells significantly increased apoptosis. High D52 but not MAL2 expression was significantly associated with reduced overall survival in breast carcinoma patients (log-rank test, P < 0.001; n = 357) and was an independent predictor of survival (hazard ratio, 2.274; 95% confidence interval, 1.228-4.210; P = 0.009; n = 328). CONCLUSION: D52 overexpression in cancer reflects specific targeting and may contribute to a more proliferative, aggressive tumor phenotype in breast cancer.
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Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Expresión Génica , Proteínas de Neoplasias/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Animales , Apoptosis/fisiología , Neoplasias de la Mama/genética , Proliferación Celular , Femenino , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Ratones , Persona de Mediana Edad , Proteínas Proteolipídicas Asociadas a Mielina y Linfocito , Proteolípidos/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Matrices Tisulares , Transfección , Proteínas de Transporte Vesicular/metabolismoRESUMEN
Simian virus 40 (SV40) is a polyomavirus for which non-human primates are the permissive host. The baboon (Papio spp.) is an old world monkey that is used in a variety of research investigations; however, natural infection of SV40 among baboons has not been thoroughly examined or reported. Initially, we were interested in determining the prevalence of SV40 infection among a captive colony of baboons based on the presence of antibodies to SV40 large T-antigen (Tag). An overall seroprevalence rate of >50% was found after screening sera from 142 baboons in the colony based on ELISA. Endpoint titer values for serum antibody binding to SV40 Tag reached as high as 1280 for 5 out of 142 baboons. Peptide binding assays revealed that a range of SV40 Tag epitopes are immunogenic in the baboon, and that individual animals differ in their humoral immune responses to SV40 Tag based on epitope recognition. Specificity to SV40 Tag and not some other primate polyomavirus encoded large Tag was further examined by serologic reactivity to peptide epitopes unique to SV40 Tag. Additional serology was performed to assess SV40 Tag reactivity by Western blot and whether antibodies were capable of neutralizing SV40 infectivity in vitro. Although antibodies with high levels of SV40 neutralization were observed in a number of the baboons, there was a lack of correlation between viral neutralization and antibodies to SV40 Tag. Further examination using molecular-based diagnosis and SV40 Tag specific real-time quantitative PCR determined that some of the baboons appeared to be exposed to SV40. DNA sequence analysis of the PCR products confirmed that SV40 Tag specific sequences were detected in baboons.
Asunto(s)
Papio/virología , Virus 40 de los Simios/aislamiento & purificación , Secuencia de Aminoácidos , Animales , Animales de Laboratorio/inmunología , Animales de Laboratorio/virología , Anticuerpos Antivirales/sangre , Antígenos Virales de Tumores/genética , Antígenos Virales de Tumores/aislamiento & purificación , Secuencia de Bases , Cartilla de ADN/genética , ADN Viral/genética , Ensayo de Inmunoadsorción Enzimática , Femenino , Masculino , Datos de Secuencia Molecular , Enfermedades de los Monos/inmunología , Enfermedades de los Monos/virología , Papio/inmunología , Papio anubis/inmunología , Papio anubis/virología , Papio cynocephalus/inmunología , Papio cynocephalus/virología , Papio ursinus/inmunología , Papio ursinus/virología , Reacción en Cadena de la Polimerasa , Infecciones por Polyomavirus/inmunología , Infecciones por Polyomavirus/veterinaria , Infecciones por Polyomavirus/virología , Homología de Secuencia de Ácido Nucleico , Estudios Seroepidemiológicos , Virus 40 de los Simios/genética , Virus 40 de los Simios/inmunología , Infecciones Tumorales por Virus/inmunología , Infecciones Tumorales por Virus/veterinaria , Infecciones Tumorales por Virus/virologíaRESUMEN
Tumor protein D52 (TPD52) is involved in transformation and metastasis and has been shown to be over-expressed in tumor cells compared to normal cells and tissues. Murine TPD52 (mD52) shares 86% protein identity with the human TPD52 orthologue (hD52). To study TPD52 protein as a target for active vaccination recombinant, mD52 was administered as a protein-based vaccine. Naïve mice were immunized with either mD52 protein and CpG/ODN as a molecular adjuvant or CpG/ODN alone. Two weeks following the final immunization, mice were challenged s.c. with syngeneic tumor cells that over-express mD52. Two distinct murine tumor cell lines were used for challenge in this model, mKSA and 3T3.mD52. Half of the mice immunized with mD52 and CpG/ODN rejected or delayed onset of mKSA s.c. tumor cell growth, and 40% of mice challenged with 3T3.mD52 rejected s.c. tumor growth, as well as the formation of spontaneous lethal lung metastases. Mice immunized with mD52 and CpG/ODN generated detectable mD52-specific IgG antibody responses indicating that mD52 protein vaccination induced an adaptive immune response. In addition, mice that rejected tumor challenge generated tumor-specific cytotoxic T lymphocytes' responses. Importantly, microscopic and gross evaluation of organs from mD52 immunized mice revealed no evidence of autoimmunity as assessed by absence of T cell infiltration and absence of microscopic pathology. Together, these data demonstrate that mD52 vaccination induces an immune response that is capable of rejecting tumors that over-express mD52 without the induction of harmful autoimmunity.