Comparative uptakes and biodistributions of internalizing vs. noninternalizing copper-64 radioimmunoconjugates in cell and animal models of colon cancer

Copper-64-labeled monoclonal antibodies (mAbs) have previously demonstrated unexpectedly effective tumor control in rodent models of cancer at relatively low tumor-absorbed radiation doses. This property has been associated with delivery platforms resulting in cellular internalization. The purpose of the present studies was to evaluate the in vitro internalization and in vivo distribution of a two-antibody model of ⁶⁴Cu radioimmunotherapy (RIT) in the same cell and animal models of cancer. Biodistributions of an internalizing antibody, cBR96, and a noninternalizing antibody, cT84.66, labeled with ⁶⁴Cu, were obtained in nude mice bearing LS174T colon carcinoma xenografts from 15 min to 48 h. The ⁶⁴Cu-DOTA-cBR96 conjugate demonstrated rapid tumor uptake, reaching 20.2% ID/g at 3 h and peaking at 35.4% ID/g by 24 h. Tumor accumulation of ⁶⁴Cu-DOTA-cT84.66 was more gradual, 8.19% ID/g at 3 h and 43.8% ID/g by 24 h, but maximum uptake was not statistically different from ⁶⁴Cu-DOTA-cBR96. Mouse xenograft dosimetry was estimated to be 1128 rad/mCi (304.9 mGy/MBq) for ⁶⁴Cu-DOTA-cBR96 and 1409 rad/mCi (380.5 mGy/MBq) for ⁶⁴Cu-DOTA-cT84.66. In LS174T cells, internalized radioactivity increased by a factor of 3.8 over 4 h for ⁶⁴Cu-DOTA-cBR96, but remained unchanged ⁶⁴Cu-DOTA-cT84.66. When normalized to uptake at 1 h, cellular efflux of ⁶⁴Cu was essentially identical for both mAbs. The biodistributions and tumor dosimetry of these internalizing and noninternalizing radiolabeled mAbs were sufficiently similar for direct comparison of the therapeutic efficacies of low doses of ⁶⁴Cu RIT agents in the same animal model of cancer.     

Escherichia coli K1 polysialic acid O-acetyltransferase gene, neuO, and the mechanism of capsule form variation involving a mobile contingency locus

Potential O-acetylation of the sialic acid residues of Escherichia coli K1, groups W-135, Y, and C meningococci, and group B Streptococcus capsular polysaccharides modifies their immunogenicity and susceptibility to glycosidases. Despite the biological importance of O-acetylation, no sialic or polysialic acid O-acetyltransferase has been identified in any system. Here we show that the E. coli K1 O-acetyltransferase encoded by neuO is genetically linked to the endo-neuraminidase tail protein gene of a chromosomal accretion element, designated CUS-3, with homology to lambdoid bacteriophage. Molecular epidemiological analysis established concordance between O-acetyltransferase and CUS-3 in a set of E. coli K1 strains. Deleting neuO eliminated enzymatic activity, which was restored by complementation in trans, and confirmed by (13)C-NMR analysis of the acetylated product. Analysis of mutants that accumulate intracellular polysialic acid because of export defects (kpsM and kpsS) or an inability to synthesize the sialic acid precursor, N-acetylmannosamine (neuC), indicated that NeuO does not require constant association with its substrate for activity. DNA sequencing and PCR analysis of neuO from strains that had undergone random capsule form variation showed that slip strand DNA mispairing or unequal recombination resulted in gain or loss of (5'-AAGACTC-3')(n) heptanucleotide repeats (where n approximately equals 14-39) located in the neuO 5' region. These repeats code for a previously undescribed structure designated the poly(Psi) motif. The unexpected discovery of the neuO contingency locus (hypervariable gene controlling expression of a surface epitope) in E. coli, and of a potential phage for redistributing variant neuO alleles, provides a robust system for investigating the functions of localized hypermutability in pathogen evolution.     

The course of cognitive functioning in first episode psychosis: changes over time and impact on outcome

This three year longitudinal study examined the cognitive performance of 247 individuals who recently presented with a first episode of psychosis. Using a comprehensive battery of cognitive tests, we assessed cognition at baseline, 1 year and 2-year follow-ups. Assessments also included positive and negative symptoms, depression, social outcome and substance use. There were several significant improvements in cognition over the two-year period which were usually matched by improvements in a matched non-psychiatric control group. Regression analyses demonstrated that after controlling for symptoms cognitive impairment accounted for 4-6% of the variance in social functioning. Our results suggest that impaired cognition exists in the very early stages of a psychotic illness and that there is no decline over time. Secondly, our results suggest that, although related, poor social functioning deficits may be independent of cognitive impairment. Finally there are implications for improved methodology in the assessment of both cognitive and social functioning.     

Behavioral analysis during the forced swimming test using a joystick device

The behavioral test described by Porsolt in 1977 for screening potential antidepressant drugs is extensively used both in basic research and in the pharmaceutical industry. The measured behavior is the immobility time during the swimming test (preformed in rodents), which decreases upon acute antidepressant treatment. Several research groups have suggested some modifications on the original Porsolt paradigm and its analysis. Nevertheless, there are still inaccuracies resulting from either undefined intermediate behaviors or from considering the movement of the whole body as one unit without analyzing the motion of the limbs. Herein, we propose a novel and simple scoring method, based on continuous measurement of the limbs motion, using a joystick, a computer screen and simple software. We validated the method, using antidepressant drugs and studied examples of false positives and false negatives of the traditional Porsolt paradigm. The proposed method is easy to use, it accounts for all range of movements and the analysis is relatively fast. Moreover, the results obtained using this analysis method show a normal Gaussian distribution in a population of rats (while the traditional Porsolt analysis does not) which allows selective breeding of 'motivated' and 'depressed' lines of animals.     

Diagnostic dilemmas in fulminant subacute sclerosing panencephalitis (SSPE)

This report describes an eleven-year-old boy with atypical features of subacute sclerosing panencephalitis (SSPE), a rare complication of measles. He had only visual symptoms for 2 months followed by rapid neurological worsening to a vegetative state in 10 days. A diagnosis of SSPE was made based on the history of measles, characteristic ocular findings, compatible magnetic resonance imaging and electroencephalographic changes, and elevated ratio of cerebrospinal fluid to serum anti-measles antibody titers     

Microcirculatory changes in patients with chronic venous and lymphatic insufficiency and heavy leg symptoms before and after therapy with procyanidol oligomers (laser-Doppler study)

INTRODUCTION: To study the efficacy of venotonica and medicaments having anti-edema effect is very actual. AIMS: Patients suffering from venous-lymphatic insufficiency were treated with procyanidol oligomers in an open, prospective study. The effective substance of this medication protects the fibrous connective tissue protein, thus improving the function of the venous and lymphatic capillaries. METHODS: For three months, the daily dose was 2 x 150 mg 30 patients were included into the study. The efficacy of the treatment was evaluated by statistical methods analysing the changes of the clinical symptoms and the satisfaction of the patients. For the first time, laser-Doppler method was used to determine the pathological condition of the dermal microcirculation before and after 3 months of treatment. The laser-Doppler tests used as indicators of the functional changing of the microcirculations were as follows: venoarterial response, reactive hyperemic response, and thermal stimulation response. The authors established the standard values and compared the measurement scores to those. According to the resulting scores, the status before and after the treatment can be compared by statistic analysis. Subjective complaints of the patients such as pain, heaviness of the leg, nightly muscle cramps were estimated by a visual analog scale. RESULTS: The treatment (procyanidol oligomers) significantly reduced lymphedema (p < 0.001) in the "heavy leg" syndrome. Symptoms as pain, heaviness of the leg, muscle cramps, improved significantly (p < 0.001) after treatment. The microcirculatory status was pathologic before treatment. After treatment, there was no significant improvement in the laser-Doppler results. CONCLUSIONS: The study revealed that in venous-lymphatic insufficiency, the arterial-capillary system is also damaged. The authors bring to attention that the disorder of the dermal arteries playing a role in the pathogenesis of venous-lymphatic insufficiency first detected in this study needs to be further investigated.     

CXCR4 regulates migration and development of human acute myelogenous leukemia stem cells in transplanted NOD/SCID mice

The chemokine stromal cell-derived factor-1 (SDF-1) and its receptor CXCR4 participate in the retention of normal hematopoietic stem cells within the bone marrow (BM) and their release into the circulation. Homing and engraftment of human stem cells in immunodeficient mice are dependent on cell surface CXCR4 expression and the production of BM SDF-1, which acts also as a survival factor for both human and murine stem cells. However, the role of SDF-1/CXCR4 interactions in the control of human acute myelogenous leukemia (AML) cell trafficking and disease progression is poorly understood. In this study, we report that although some AML cells do not express surface CXCR4, all AML cells tested express internal CXCR4 and SDF-1. Culture of AML cells with SDF-1 promoted their survival, whereas addition of neutralizing CXCR4 antibodies, SDF-1 antibodies, or AMD3100 significantly decreased it. Pretreatment of primary human AML cells with neutralizing CXCR4 antibodies blocked their homing into the BM and spleen of transplanted NOD/SCID/B2m(null) mice. Furthermore, weekly administrations of antihuman CXCR4 to mice previously engrafted with primary AML cells led to a dramatic decrease in the levels of human AML cells in the BM, blood, and spleen in a dose- and time-dependent manner. Interestingly, the same treatment did not affect significantly the levels of normal human progenitors engrafted into NOD/SCID mice. Taken together, our findings demonstrated the importance of the SDF-1/CXCR4 axis in the regulation of in vivo motility and development of human AML stem cells and identified CXCR4 neutralization as a potential treatment for AML.