The p53 gene product is overexpressed in approximately 50% of cancers, making it an ideal target for cancer immunotherapy. We previously demonstrated that a modified vaccinia Ankara (MVA) vaccine ...expressing human p53 (MVA-p53) was moderately active when given as a homologous prime/boost in a human p53 knock in (Hupki) mouse model. We needed to improve upon the inefficient homologous boosting approach, because development of neutralizing immunity to the vaccine viral vector backbone suppresses its immunogenicity. To enhance specificity, we examined the combination of 2 different vaccine vectors provided in sequence as a heterologous prime/boost. Hupki mice were evaluated as a human p53 tolerant model to explore the capacity of heterologous p53 immunization to reject human p53-expressing tumors. We employed attenuated recombinant Listeria monocytogenes expressing human p53 (LmddA-LLO-p53) in addition to MVA-p53. Heterologous p53 immunization resulted in a significant increase in p53-specific CD8 and CD4 T cells compared with homologous single vector p53 immunization. Heterologous p53 immunization induced protection against tumor growth but had only a modest effect on established tumors. To enhance the immune response we used synthetic double-strand RNA (polyinsosinic:polycytidylic acid) and unmethylated CpG-containing oligodeoxynucleotide to activate the innate immune system via Toll-like receptors. Treatment of established tumor-bearing Hupki mice with polyinsosinic:polycytidylic acid and CpG-oligodeoxynucleotide in combination with heterologous p53 immunization resulted in enhanced tumor rejection relative to treatment with either agent alone. These results suggest that heterologous prime/boost immunization and Toll-like receptor stimulation increases the efficacy of a cancer vaccine, targeting a tolerized tumor antigen.
Background Genetically triggered thoracic aortic conditions (GenTACs) represent an important problem for patients and their families. Accordingly, the National Heart, Lung, and Blood Institute ...established the first phase of its national GenTAC Registry in 2006. Enrollment and Diagnoses Between 2007 and 2010, 6 enrolling centers established the GenTAC I Registry consisting of 2,046 patients (Marfan syndrome 576 28.2%, bicuspid aortic valve disease 504 24.6%, aneurysm or dissection age <50 years 369 18%, and others). Biologic samples for DNA analyses (white blood cells or saliva) are available in 97%, and stored plasma is available in 60% of enrollees. Results Initial scientific inquiry using the GenTAC Registry has included validation studies of genetic causes for aortic syndromes, potential usefulness of transforming growth factor beta (TGFB) blood levels in Marfan subjects, and current surgical approaches to ascending aortic conditions. Future Opportunity The second phase of GenTAC will allow biannual follow-up of GenTAC I enrollees for up to 9 years, enrollment of an additional 1,500 subjects, further integration of imaging findings with clinical and genetic data through utilization of an imaging core laboratory, important validation of phenotype-genotype correlations through a phenotyping core laboratory, and integration of a scientific advisory committee to help define the full range and depth of the Registry's scientific capabilities. The registry resources are available to the external scientific community through an application process accessible at https://gentac.rti.org.
Retinoids elicit biological responses by activating a series of nuclear receptors. Six retinoid receptors belonging to two families are currently known: retinoic acid receptors (RAR alpha,beta,and ...gamma) and retinoid X receptors (RXR alpha,beta,and gamma). Stilbene retinoid analogs of retinoic acid (RA), such as (E)-4-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)prope n-1- ylbenzoic acid (TTNPB, 1) and (E)-4-2-(5,6,7,8-tetrahydro-3,5,5,8,8-pentamethyl-2-naphthalenyl)pro pen-1- ylbenzoic acid (3-methyl-TTNPB, 2), display differential RAR and RXR activities, depending on the substituent at C3 of the naphthalene ring. We report here structural modifications of the benzoate moiety of 2 that result in analogs with greater RXR selectivity as well as those with pan-agonist (activate both RAR and RXR receptors) activities, analyze the structural features that impart receptor selectivity, and describe a stereoselective method for the synthesis of these analogs. The biological activities associated with the RAR and RXR receptors were examined by testing representative examples with different receptor activation profiles for their ability to induce tissue transglutaminase (Tgase) activity in a human promyelocytic leukemia cell line (HL-60 cdm-1) and to inhibit tumor-promoter-induced ornithine decarboxylase (ODC) activity in hairless mouse skin. These results suggest that RAR agonists and RXR agonists may have different therapeutic applications. Finally, we show that RXR agonists are significantly reduced in teratogenic potency relative to RAR agonists and may therefore have significant advantages in clinical practice.
Background Several technologies have been developed to aid dermatologists in the detection of melanoma in vivo including dermoscopy, multispectral digital skin lesion analysis (MDSLA), and ...reflectance confocal microscopy (RCM). To our knowledge, there have been no studies directly comparing MDSLA and RCM. Objective We conducted a repeated measures analysis comparing the sensitivity and specificity of MDSLA and RCM in the detection of melanoma (n = 55 lesions from 36 patients). Methods Study patients (n = 36) with atypical-appearing pigmented lesions (n = 55) underwent imaging by both RCM and MDSLA. Lesions were biopsied and analyzed by histopathology. Results RCM exhibited superior test metrics ( P = .001, McNemar test) compared with MDSLA. Respectively, sensitivity measures were 85.7% and 71.4%, and specificity rates were 66.7% and 25.0%. Limitations The sample size was relatively small and was collected from only one dermatologist's patient base; there was some degree of dermatopathologist interobserver variability; and only one confocalist performed the RCM image evaluations. Conclusion RCM is a useful adjunct during clinical assessment of in vivo lesions suspicious for melanoma or those requiring re-excision because of high level of dysplasia or having features consistent with an atypical melanocytic nevus with severe cytologic atypia.
Serological analysis of cDNA expression library (SEREX) was employed to identify cancer-associated genes. By screening cDNA expression libraries with sera of patients with lung cancers, we identified ...a total of 49 genes that specifically reacted with the sera of patients with lung cancers. Among these, we characterized a novel gene with expression pattern similar to that of cancer/testis antigens. Its open reading frame is 1920 bp in size and encodes for putative protein composed of 639 amino acids. Southern blot analysis reveals that this gene exists as single copy. In vitro transcription/translation and Western blot analysis confirm that this gene encodes a protein of 73 kDa in size. The comparison of cDNA and genomic sequences reveals that it is composed of 11 exons and 10 introns. This gene displays testis-specific expression among normal tissues, and wide expression among various cancer tissues and cancer cell lines. A study using GFP fusion construct reveals mainly nuclear localization of CAGE-1 protein. The expression of this clone is relatively higher in cancer tissues compared with their surrounding non-cancerous tissues. This suggests that overexpression of CAGE-1 may be associated with the progression of tumor. Because of its association with cancer, this gene was named cancer-associated gene-1 (CAGE-1). Given the fact that several cancer/testis antigens reportedly induce cytolytic T lymphocyte (CTL) reactions, it is reasonable that this gene will be a valuable target for cancer immunotherapy. The exact functional role of CAGE-1 in tumorigenesis remains to be seen.
The effect of antiserum against Met
5-enkephalin, Leu
5-enkephalin, β-endorphin, or dynorphin A-(1-13) administered intracerebroventricularly (i.c.v.) or intrathecally (i.t.) on ...immobilization-induced antinociception was studied in ICR mice. Antinociception was assessed by the tail-flick assay. Immobilization of the mouse increased inhibition of the tail-flick response at least 1 h. The i.c.v. or i.t. injection with antiserum against dynorphin A-(1-13) at the dose of 200 μg significantly attenuated immobilization-induced inhibition of the tail-flick response. However, antiserum against Met
5-enkephalin, Leu
5-enkephalin, or β-endorphin did not affect the immobilization stress-induced antinociception. Furthermore, i.c.v. or i.t. injection with nor-binaltorphimine (Nor-BNI; from 1 to 20 μg) effectively inhibited immobilization stress-induced inhibition of the tail-flick response in a dose-dependent manner. However, β-FNA (from 0.5 to 2 μg) or naltrindole (from 1 to 20 μg) administered i.c.v. or i.t. did not affect immobilization stress-induced antinociception. Our results suggest that supraspinally and spinally located dynorphin appears to be involved in the production of immobilization stress-induced antinociception via stimulating κ-opioid receptors.
A 116-kDa polypeptide has recently been found to be a common component of vacuolar proton pumps isolated from a variety of sources. The 116-kDa subunit of the proton pump was purified from ...clathrin-coated vesicles of bovine brain, and internal sequences were obtained from proteolytic peptides. Oligonucleotide probes designed from these peptide sequences were utilized in polymerase chain reactions to isolate partial bovine cDNA clones for the protein. Sequences from these were then utilized to isolate rat brain cDNA clones containing the full-length coding region. RNA blots indicate the presence of an abundant 3.9-kilobase message for the 116-kDa subunit in brain, and primer extension analysis demonstrates that the cloned sequence is full-length. The rat cDNA sequences predict synthesis of a protein of 96,267 Da. Analysis of the deduced amino acid sequence of the 116-kDa subunit suggests that it consists of two fundamental domains: a hydrophilic amino-terminal half that is composed of greater than 30% charged residues, and a hydrophobic carboxyl-terminal half that contains at least six transmembrane regions. The structural properties of the 116-kDa proton pump polypeptide agree well with its proposed function in coupling ATP hydrolysis by the cytoplasmic subunits to proton translocation by the intramembranous components of the pump.
We studied whether eugenol provides neuroprotection against delayed neuronal death in the hippocampal CA1 region following a 5 min occlusion of the common carotid arteries bilaterally under either ...free-regulating temperature (TF) or maintained temperature (TM, 37ºC) conditions in gerbils. Right after occlusion of the carotid arteries, we injected eugenol intraperitoneally at concentrations of either 50, 100, or 200 mg/kg. There was significant preservation of neuronal cells in the CA1 region in the eugenol-treated groups 7 days after the ischemic insult in the TF condition, with respective survival values of 26, 43, and 68%. In the TM condition, however, significant neuroprotection was only seen with eugenol concentrations of 100 and 200 mg/kg (32% and 52%, respectively). When the rectal temperature was maintained at 38ºC for 30 min after occlusion of the carotid arteries, no reduction in CA1 damage was observed with any dose of eugenol. These results suggest that eugenol may provide neuroprotection against ischemic damage by its hypothermic action.
Background There are currently 2 existing confocal laser endomicroscopy (CLE) platforms: probe-based CLE (pCLE) and endoscope-based CLE (eCLE) systems, each with its own criteria for identifying ...dysplasia in Barrett's esophagus (BE). The diagnostic performance of these 2 systems has not been directly compared. Design Preclinical, feasibility study. Objectives We compared the interrater agreement and diagnostic performance of the pCLE and eCLE systems. In addition, we evaluated a new BE endomicroscopy criteria based on fluorescent glucose intensity uptake. Patients Thirteen patients with Barrett's esophagus and high-grade dysplasia or early cancer undergoing 16 EMR. Intervention CLE imaging was performed using two different probes with 2-N-(7-nitrobenz-2-oxa-1,3-diaxol-4-yl)amino-2-deoxyglucose, a fluorescent glucose analog with preferential uptake in dysplastic mucosa to supply contrast. Four quadrants were imaged per specimen with a total of 64 imaged mucosal sites presented to three gastroenterologists. Main Outcome Measurements Interobserver agreement and accuracy for dysplasia was assessed of images classified according to Miami criteria, stacked eCLE images classified using the Mainz criteria and a novel fluorescence intensity criteria. Results The interrater agreements were 0.17, 0.68, and 0.87 for the Miami, Mainz, and the fluorescence intensity criteria, respectively. Overall accuracy in detecting dysplasia was 37% (95% CI, 30.3-43.9), 44.3% (95% CI, 37.3-50.9), and 78.6% (95% CI, 72.2-83.3) for the Miami, Mainz, and the fluorescence intensity criteria, respectively. Limitations This imaging technique and proposed fluorescence intensity criteria using 2-N-(7-nitrobenz-2-oxa-1,3-diaxol-4-yl)amino-2-deoxyglucose in EMR tissue will require in vivo validation and cannot be directly used with the current eCLE and pCLE clinical applications. Conclusions In this preclinical feasibility study, the use of an eCLE system with a topical fluorescent contrast in ex vivo EMR tissue demonstrated higher interrater agreement and accuracy.
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