Citations

The National Heart, Lung, and Blood Institute (NHLBI) convened a conference call working group, consisting of experts in stroke and cerebrovascular biology on January 28, 2005. The purpose of this working group was to develop a prioritized set of recommendations for NHLBI to establish a focused and comprehensive set of research activities in cerebrovascular biology and disease. Three thematic areas of research emerged: (1) molecular and cellular neurobiology of cerebral blood vessels, focusing on genomics and proteomics, neurovascular signaling and cerebrovascular embryogenesis, development and plasticity; (2) resource development, involving the development of new methodological approaches for normal and altered function of the neurovascular unit, collaborative research, and training in cerebrovascular pathobiology; and (3) cerebrovascular diseases and translational approaches, addressing vascular mechanisms of disease, the role of risk factors, importance of biomarkers with the ultimate goal of developing new treatments.

Background: An uncontrolled pilot study demonstrated that daclizumab, a humanised monoclonal antibody to the interleukin 2 receptor (CD25), might be effective for the treatment of active ulcerative colitis. Methods: A randomised, double blind, placebo controlled trial was conducted to evaluate the efficacy of daclizumab induction therapy in patients with active ulcerative colitis. A total of 159 patients with moderate ulcerative colitis were randomised to receive induction therapy with daclizumab 1 mg/kg intravenously at weeks 0 and 4, or 2 mg/kg intravenously at weeks 0, 2, 4, and 6, or placebo. The primary end point was induction of remission at week 8. Remission was defined as a Mayo score of 0 on both endoscopy and rectal bleeding components and a score of 0 or 1 on stool frequency and physician’s global assessment components. Response was defined as a decrease from baseline in the Mayo score of at least 3 points. Results: Two per cent of patients receiving daclizumab 1 mg/kg (p = 0.11 v placebo) and 7% of patients receiving 2 mg/kg (p = 0.73) were in remission at week 8, compared with 10% of those who received placebo. Response occurred at week 8 in 25% of patients receiving daclizumab 1 mg/kg (p = 0.04) and in 33% of patients receiving 2 mg/kg (p = 0.30) versus 44% of those receiving placebo. Daclizumab was well tolerated. The most frequently reported adverse events in daclizumab treated patients compared with placebo treated patients were nasopharyngitis (14.6%) and pyrexia (10.7%). Conclusion: Patients with moderate ulcerative colitis who are treated with daclizumab are not more likely to be in remission or response at eight weeks than patients treated with placebo.

Recent studies report that, in the absence of heart failure and renal failure, plasma B-type natriuretic peptide (BNP) has prognostic value for mortality. We sought to confirm and extend these previous studies to assess BNP, measured by 3 distinct assays, as a biomarker for mortality in a strategy to enhance efforts at primary prevention and to better understand the clinical phenotype of such subjects at risk. We used a community-based cohort of 2042 subjects from Olmsted County, Minn, and individuals with heart or renal failure were excluded. BNP was assessed using 3 assays including Biosite and Shionogi for mature, biologically active BNP and the Roche assay for apparently nonbiologically active amino-terminal pro-BNP (NT-proBNP). Thorough echocardiographic and clinical data were recorded for all of the participants. Median follow-up for mortality was 5.6 years. BNP by all 3 of the assays was predictive of mortality. NT-proBNP and Biosite assays remained significant even after adjustment for traditional clinical risk factors and echocardiographic abnormalities including left ventricular hypertrophy and diastolic dysfunction. Echocardiography documented widespread structural changes in those with increasing BNP levels yet below levels observed in heart failure. We report in a large, well-characterized community-based cohort, free of heart failure, the first study to compare 3 distinct BNP assays as biomarkers for mortality in the same cohort. Our findings confirm the potential use of NT-proBNP and BNP biomarkers for future events and underscore that these peptides may also serve as biomarkers for underlying cardiac remodeling secondary to diverse cardiovascular disease entities.

Abstract Purpose: Gemcitabine is a nucleoside analogue with activity against solid tumors. Gemcitabine metabolic inactivation is catalyzed by cytidine deaminase (CDA) or, after phosphorylation, by deoxycytidylate deaminase (DCTD). We set out to study the pharmacogenomics of CDA and DCTD. Experimental Design: The genes encoding CDA and DCTD were resequenced using DNA from 60 African American and 60 Caucasian American subjects. Expression constructs were created for nonsynonymous coding single nucleotide polymorphisms (cSNP) and reporter gene constructs were created for 5′-flanking region polymorphisms. Functional genomic studies were then conducted after the transfection of mammalian cells. Results: CDA resequencing revealed 17 polymorphisms, including one common nonsynonymous cSNP, 79 A>C (Lys27Gln). Recombinant Gln27 CDA had 66 ± 5.1% (mean ± SE) of the wild-type (WT) activity for gemcitabine but without a significant decrease in level of immunoreactive protein. The apparent Km (397 ± 40 μmol/L) for the Gln27 allozyme was significantly higher than that for the WT (289 ± 20 μmol/L; P < 0.025). CDA 5′-flanking region reporter gene studies showed significant differences among 5′-flanking region haplotypes in their ability to drive transcription. There were 29 SNPs in DCTD, including one nonsynonymous cSNP, 172 A>G (Asn58Asp), in Caucasian American DNA. Recombinant Asp58 DCTD had 11 ± 1.4% of WT activity for gemcitabine monophosphate with a significantly elevated level of immunoreactive protein. No DCTD polymorphisms were observed in the initial 500 bp of the 5′-flanking region. Conclusions: These results suggest that pharmacogenomic variation in the deamination of gemcitabine and its monophosphate might contribute to variation in therapeutic response to this antineoplastic agent.

Antibodies are generally thought to be a class of proteins that function without the use of cofactors. However, it is not widely appreciated that antibodies are believed to be the major carrier protein in human circulation for the important riboflavin cofactor that is involved in a host of biological phenomena. A further link between riboflavin and antibodies was discovered 30 years ago when a bright-yellow antibody, IgG GAR , was purified from a patient with multiple myeloma who had turned yellow during the course of her disease. It was subsequently shown that the yellow color of this antibody was due to riboflavin binding. However, it was not known how and where riboflavin was bound to this antibody. We now report the crystal structure of this historically important IgG GAR Fab at 3.0-Å resolution. The riboflavin is located in the antigen-combining site with its isoalloxazine ring stacked between the parallel aromatic moieties of TyrH33, PheH58, and TyrH100A. Together with additional hydrogen bonds, these interactions reveal the structural basis for high-affinity riboflavin binding. The ligand specificity of IgG GAR is compared with another riboflavin-binding antibody, IgG DOT , which was purified from a second patient with multiple myeloma. The crystal structure of IgG GAR provides a starting point for attempts to understand the physiological relevance and chemical functions of cofactor-containing antibodies.

Abstract The ability of cancers to evade immune surveillance and resist immunotherapy raises a fundamental question of how tumor cells survive in the presence of a competent immune system. Studies to address this question have primarily focused on mechanisms by which tumor cells avoid recognition by or induce tolerance in the immune system. However, little is known about whether cancer cells also acquire an intrinsic ability to resist killing by immune effectors. We find that cancer cells enhance their ability to withstand an attack by cytotoxic immune effector cells via acquisition of specific genetic alterations that interfere with the shared mitochondrial death signaling pathway entrained by granzyme B, IFN-γ, and Apo2 ligand/tumor necrosis factor–related apoptosis inducing ligand (Apo2L/TRAIL), three key mediators of immunologic cell–mediated cytotoxicity. We show that the coexistence of specific mitochondrial signaling defects (either deletion of Bax, overexpression of Bcl-xL, or deletion of Smac) with expression of X-linked inhibitor of apoptosis protein decreases the sensitivity of cancer cells to IFN-γ/Apo2L/TRAIL– or granzyme B–induced apoptosis, lymphocyte-mediated cytotoxicity in vitro, and adoptive cellular immunotherapy in vivo. Conversely, negating X-linked inhibitor of apoptosis protein expression or function in tumor cells with defective mitochondrial signaling enables direct activation of caspase-3/-7 by granzyme B or Apo2L/TRAIL, and restores their susceptibility to immunologic cytotoxicity. These findings identify an important mechanism by which cancers evade elimination by immune effector cells and suggest that cancer immunotherapy might be improved by concurrent strategies to alleviate or circumvent the intrinsic mitochondrial death signaling defects that help cancer cells resist immunologic cytotoxicity. (Cancer Res 2006; 66(3): 1730-9)

Abstract Lapatinib (GW572016) is a selective inhibitor of both epidermal growth factor receptor (EGFR) and HER-2 tyrosine kinases. Here, we explore the therapeutic potential of lapatinib by testing its effect on tumor cell growth in a panel of 31 characterized human breast cancer cell lines, including trastuzumab-conditioned HER-2-positive cell lines. We further characterize its activity in combination with trastuzumab and analyze whether EGFR and HER-2 expression or changes induced in the activation of EGFR, HER-2, Raf, AKT, or extracellular signal-regulated kinase (ERK) are markers of drug activity. We report that concentration-dependent antiproliferative effects of lapatinib were seen in all breast cancer cell lines tested but varied significantly between individual cell lines with up to 1,000-fold difference in the IC50s (range, 0.010-18.6 μmol/L). Response to lapatinib was significantly correlated with HER-2 expression and its ability to inhibit HER-2, Raf, AKT, and ERK phosphorylation. Long-term in vivo lapatinib studies were conducted with human breast cancer xenografts in athymic mice. Treatment over 77 days resulted in a sustained and significant reduction in xenograft volume compared with untreated controls. For the combination of lapatinib plus trastuzumab, synergistic drug interactions were observed in four different HER-2-overexpressing cell lines. Moreover, lapatinib retained significant in vitro activity against cell lines selected for long-term outgrowth (>9 months) in trastuzumab-containing (100 μg/mL) culture medium. These observations provide a clear biological rationale to test lapatinib as a single agent or in combination with trastuzumab in HER-2-overexpressing breast cancer and in patients with clinical resistance to trastuzumab. (Cancer Res 2006; 66(3): 1630-9)