The DIRE score: predicting outcomes of opioid prescribing for chronic pain.

The objective of this retrospective study was to test the validity and reliability of a scoring tool (the DIRE Score), for use by clinicians, that predicts which chronic noncancer pain patients will have effective analgesia and be compliant with long-term opioid maintenance treatment. DIRE scores were assigned to 61 cases from the pain center's databases. These cases were abstracted into vignettes that were reviewed and scored by 6 physicians. Repeat scoring was carried out on a subset of 30 vignettes after 2 weeks. The main outcome measures were: global impression of compliance and efficacy as indicated in the medical record and by interview with the patient's treating clinician; and final disposition, ie, whether or not opioids were continued or discontinued at the time of last clinical documentation. Internal consistency of the factors making up the DIRE Score was high (Cronbach's alpha = .80). Sensitivity and specificity of the DIRE Score for predicting patient compliance were 94% and 87%, respectively. For efficacy, sensitivity and specificity were 81% and 76%. For disposition, the sensitivity and specificity were 86% and 73%. Intraclass correlation was 0.94 for interrater reliability and 0.95 for intrarater reliability. PERSPECTIVE: Public controversy about the use of long-term opioids for chronic pain fuels physician ambivalence about the prescribing process. In this initial retrospective study, validity and reliability of the DIRE Score are demonstrated. The score correlated well with measures of patient compliance and efficacy of long-term opioid therapy.     

Differential effects of delta9-tetrahydrocannabinol and methanandamide in CB1 knockout and wild-type mice

Mice devoid of CB1 cannabinoid receptors (CB1-/- mice) provide a unique opportunity to further investigate the role of CB1 receptors in exocannabinoid and endocannabinoid effects. CB1-/- mice (N = 18) and their wild-type littermates (CB1+/+ mice; N = 12) were placed in standard mouse operant chambers and trained to lever press under a fixed ratio 10 schedule of reinforcement. When stable lever press responding under the fixed ratio 10 schedule had been established, cannabinoids and noncannabinoids were administered to both groups. CB1+/+ mice acquired the lever press response more readily than CB1-/- mice. Delta(9)-Tetrahydrocannabinol (Delta(9)-THC) decreased lever press responding in CB1+/+ mice only, whereas methanandamide, a metabolically stable endocannabinoid analog, produced similar response rate decreases in both genotypic groups. Similar to Delta(9)-THC, another endocannabinoid analog, (R)-(20-cyano-16,16-dimethyl docosa-cis-5,8,11,14-tetraeno)-1'-hydroxy-2'-propylamine (O-1812), decreased responding in CB1+/+ mice, but not in CB1-/- mice. The CB1 receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl-4-methyl-1H-pyrazole-3-carboxamide hydrochloride (SR141716A) blocked the effects of Delta(9)-THC, but not those of methanandamide. Because methanandamide binds poorly to CB2 receptors, these results suggest possible non-CB1, non-CB2 mechanisms of action for methanandamide-induced behavioral disruption of lever press responding. Ethanol and morphine elicited greater response decreases in CB1-/- mice than in CB1+/+ mice, suggesting a possible role of CB1 receptors in the rate disruptive effects of these drugs. In contrast, diazepam did not produce between group differences, suggesting that CB1 receptors are not involved in diazepam-induced disruption of lever press responding.     

Age at Regular Drinking,Clinical Course,and Heritability of Alcohol Dependence in the San Francisco Family Study: A Gender Analysis

We examined gender differences in age of onset, clinical course, and heritability of alcohol dependence in 2,524 adults participating in the University of California San Francisco (UCSF) family study of alcoholism. Men were significantly more likely than women to have initiated regular drinking during adolescence. Onset of regular drinking was not found to be heritable but was found to be significantly associated with a shorter time to onset of alcohol dependence. A high degree of similarity in the sequence of alcohol‐related life events was found between men and women, however, men experienced alcohol dependence symptoms at a younger age and women had a more rapid clinical course. Women were found to have a higher heritability estimate for alcohol dependence (h²= .46) than men (h²= .32). These findings suggest that environmental factors influencing the initiation of regular drinking rather than genetic factors associated with dependence may in part underlie some of the gender differences seen in the prevalence of alcohol dependence in this population. (Am J Addict 2010;00:1–10)     

Formation mechanism and structure of a guanine-uracil DNA intrastrand cross-link

The formation and structure of the 5'-G[8-5]U-3' intrastrand cross-link are studied using density functional theory and molecular dynamics simulations due to the potential role of this lesion in the activity of 5-halouracils in antitumor therapies. Upon UV irradiation of 5-halouracil-containing DNA, a guanine radical cation reacts with the uracil radical to form the cross-link, which involves phosphorescence or an intersystem crossing and a rate-determining step of bond formation. Following ionizing radiation, guanine and the uracil radical react, with a rate-limiting step involving hydrogen atom removal. Although cross-link formation from UV radiation is favored, comparison of calculated reaction thermokinetics with that for related experimentally observed purine-pyrimidine cross-links suggests this lesion is also likely to form from ionizing radiation. For the first time, the structure of 5'-G[8-5]U-3' within DNA is identified by molecular dynamics simulations. Furthermore, three conformations of cross-linked DNA are revealed, which differ in the configuration of the complementary bases. Distortions, such as unwinding, are localized to the cross-linked dinucleotide and complementary nucleotides, with minimal changes to the flanking bases. Global changes to the helix, such as bending and groove alterations, parallel cisplatin-induced distortions, which indicate 5'-G[8-5]U-3', may contribute to the cytotoxicity of halouracils in tumor cell DNA using similar mechanisms.     

Positive traits versus previous trauma: racially different correlates with PTSD symptoms among hurricane katrina‐rita volunteers

This study compared risks and protective factors for acquiring symptoms of posttraumatic stress disorder (PTSD) between African‐American (n=299) and European‐American (n=206) student volunteers 3 months after Hurricanes Katrina and Rita (H‐KR). Respondents retrospectively provided information on peritraumatic emotional reactions and previous trauma that were recalled by H‐KR and H‐KR stressors. African‐American respondents reported higher levels of symptoms and higher rates of recollection of prior traumas during H‐KR than their European‐American counterparts. Hierarchical regression analyses found that previous trauma recollections predicted symptoms among European Americans but not among African Americans. Disaster‐related stressors, however, affected African Americans more than European Americans. Though negative emotions had negative outcomes for both groups, positive emotions and hope served as protective factors for African Americans. © 2011 Wiley Periodicals, Inc.     

Anti‐retroviral effects of type I IFN subtypes in vivo

Type I IFN play a very important role in immunity against viral infections. Murine type I IFN belongs to a multigene family including 14 IFN‐α subtypes but the biological functions of IFN‐α subtypes in retroviral infections are unknown. We have used the Friend retrovirus model to determine the anti‐viral effects of IFN‐α subtypes in vitro and in vivo. IFN‐α subtypes α1, α4, α6 or α9 suppressed Friend virus (FV) replication in vitro, but differed greatly in their anti‐viral efficacy in vivo. Treatment of FV‐infected mice with the IFN‐α subtypes α1, α4 or α9, but not α6 led to a significant reduction in viral loads. Decreased splenic viral load after IFN‐α1 treatment correlated with an expansion of activated FV‐specific CD8⁺ T cells and NK cells into the spleen, whereas in IFN‐α4‐ and ‐α9‐treated mice it exclusively correlated with the activation of NK cells. The results demonstrate the distinct anti‐retroviral effects of different IFN‐α subtypes, which may be relevant for new therapeutic approaches.     

Neutralizing Monoclonal Antibodies Directed against Defined Linear Epitopes on Domain 4 of Anthrax Protective Antigen

The anthrax protective antigen (PA) is the receptor-binding subunit common to lethal toxin (LT) and edema toxin (ET), which are responsible for the high mortality rates associated with inhalational Bacillus anthracis infection. Although recombinant PA (rPA) is likely to be an important constituent of any future anthrax vaccine, evaluation of the efficacies of the various candidate rPA vaccines is currently difficult, because the specific B-cell epitopes involved in toxin neutralization have not been completely defined. In this study, we describe the identification and characterization of two murine monoclonal immunoglobulin G1 antibodies (MAbs), 1-F1 and 2-B12, which recognize distinct linear neutralizing epitopes on domain 4 of PA. 1-F1 recognized a 12-mer peptide corresponding to residues 692 to 703; this epitope maps to a region of domain 4 known to interact with the anthrax toxin receptor CMG-2 and within a conformation-dependent epitope recognized by the well-characterized neutralizing MAb 14B7. As expected, 1-F1 blocked PA''s ability to associate with CMG-2 in an in vitro solid-phase binding assay, and it protected murine macrophage cells from intoxication with LT. 2-B12 recognized a 12-mer peptide corresponding to residues 716 to 727, an epitope located immediately adjacent to the core 14B7 binding site and a stretch of amino acids not previously identified as a target of neutralizing antibodies. 2-B12 was as effective as 1-F1 in neutralizing LT in vitro, although it only partially inhibited PA binding to its receptor. Mice passively administered 1-F1 or 2-B12 were partially protected against a lethal challenge with LT. These results advance our fundamental understanding of the mechanisms by which antibodies neutralize anthrax toxin and may have future application in the evaluation of candidate rPA vaccines.Bacillus anthracis has long been recognized as a serious public health threat, given the ease with which B. anthracis spores can be disseminated via aerosol and due to the high mortality rate that accompanies spore inhalation. These fears were realized in the fall of 2001, when B. anthracis spores were circulated through the U.S. postal system, resulting in five deaths, 22 known cases of infection, and the possible exposure of more than 30,000 people (7). Although routine vaccination of civilians against anthrax is neither necessary nor desirable, certain segments of the population, notably emergency first responders and research laboratory personnel, remain at risk of exposure and are in need of an effective preexposure vaccine. In the United States, the only licensed anthrax vaccine, Anthrax Vaccine Adsorbed (AVA) or Biothrax, has been subject to controversy for years, due to safety concerns (3), and is not considered an ideal vaccine due to its protracted vaccination schedule (six injections over 18 months). AVA, which consists of formalin-treated culture filtrate from an attenuated strain of B. anthracis adsorbed to aluminum hydroxide, is also inherently difficult to manufacture and to standardize. A recombinant anthrax vaccine, manufactured by VaxGen, was considered to be a leading candidate to replace AVA and to supply the Strategic National Stockpile with 75 million doses, but it was recently discontinued because of concerns regarding vaccine production and stability. For these reasons, there is now a concerted effort to develop a new anthrax vaccine based on a defined antigenic composition.Current anthrax vaccines are focused on protective antigen (PA), the primary determinant in AVA that is involved in immunity to anthrax infection (21). PA is the receptor-binding subunit common to both lethal factor (LF), and edema factor (EF), which, upon PA binding form lethal toxin (LT) and edema toxin (ET), respectively. The anthrax toxins are responsible for the high mortality rates associated with inhalational B. anthracis infection. PA is secreted by B. anthracis as an 83-kDa protein (PA₈₃) that binds two known anthrax toxin receptors (ATRs), namely, tumor endothelium marker 8 (TEM-8) and capillary morphogenesis protein 2 (CMG-2) (5, 6, 19). Following receptor engagement, the amino-terminal 20-kDa region of the PA (PA₂₀) is proteolytically cleaved by a furinlike protease, exposing a region of PA that allows for the formation of the holotoxin. The membrane-bound 63-kDa PA (PA₆₃) spontaneously oligomerizes into heptamers and then associates either with LF, a zinc-dependent protease specific for certain members of the mitogen-activated protein kinase kinase family (12, 13), or EF, a calmodulin-, Ca²⁺-dependent adenylate cyclase (20), to form LT and ET.It is well established that animals immunized with recombinant PA, or passively administered anti-PA antiserum, are protected against LT/ET exposure or spore challenge (reviewed in references 16 and 7). For example, we recently reported that LT-challenged mice treated with goat anti-PA antiserum (either alone or in combination with antibiotics) demonstrated increased survival rates compared to untreated mice (18). However, the antibody response to PA is complicated: total anti-PA titers, as measured by enzyme-linked immunosorbent assays (ELISAs), do not correlate well with protection (35). This is likely due to the fact that a large number of anti-PA antibodies are directed against non-neutralizing (or possibly even toxin-enhancing) epitopes (29, 41). Pioneering work by Little et al. (23) suggests that at least 20 antigenic determinants exist on PA. At present, only a few of these epitopes has been described in detail; several have been shown to be targets of monoclonal antibodies (MAbs) capable of effective neutralization in vivo or in vitro (1, 8, 10, 17, 44, 45). The fact that key neutralizing epitopes on PA remain widely unidentified poses a significant barrier to the evaluation of the efficacy of vaccines based on full-length and truncated PA subunits.PAs can be divided into four functional domains. Domain 1 (residues 1 to 258) contains the furin recognition site (¹⁶⁴RKKR¹⁶⁷), as well as the region of PA that is recognized by LF and EF (30). Domain 2 (residues 259 to 487) and part of domain 3 (residues 488 to 595) are implicated in heptamerization, pore formation, and translocation of EF/LF across endosomal membranes (4, 27, 28, 34). Domain 4 (residues 596 to 735) constitutes the region of the PA involved in receptor recognition and attachment (39). Antibodies against domain 4 are postulated to be the most effective in neutralizing LT and ET (1), since they are proposed to interfere with PA binding to ATR. However, the only neutralizing epitope that has been characterized in detail is the one recognized by MAb 14B7 (24, 37, 40). This MAb contacts the face of domain 4 that is involved in receptor recognition, and has been proposed to encompass residues 671 to 721. Certainly, additional neutralizing epitopes exist on domain 4 (1, 46). In particular, Abboud and Casadevall (1) suggested a linear epitope, immediately adjacent to or possibly overlapping the 14B7 binding site, as the target of neutralizing antibodies. The characterization of antibodies that are both linear epitope dependent and neutralizing will be useful in the evaluation specific correlates of immunity; such MAbs can be incorporated into functional screening assays of potential vaccine candidates.The goal of our study was to identify additional B-cell epitopes, within domain 4 of PA, that constitute the targets of neutralizing MAbs. Toward this end, we screened a collection of B-cell hybridomas, produced from PA₈₃-immunized mice, for MAbs capable of binding to a domain 4-specific peptide array. We identified two domain 4-reactive MAbs, 1-F1 and 2-B12. 1-F1 recognized a peptide spanning residues 692 to 703; this epitope maps to a region of domain 4 that overlaps with the binding site recognized by the well-characterized neutralizing antibody 14B7 (residues 671 to 721) (23, 40). 2-B12, on the other hand, recognized a peptide spanning residues 716 to 727; this epitope maps adjacent to the core 14B7 binding site and at a location not previously identified as being a target of neutralizing antibodies. Both MAbs neutralized LT in vitro and partially protected mice against LT challenge. These results advance our fundamental understanding of the mechanisms by which antibodies neutralize anthrax toxin and may have future application in the evaluation of candidate rPA vaccines.