Enzyme assisted routes to bioactive molecules

981-99

GD2-directed bispecific trifunctional antibody outperforms dinutuximab beta in a murine model for aggressive metastasized neuroblastoma

Background: Neuroblastoma is the most common extracranial solid tumor of childhood. Patients with high-risk disease undergo extremely aggressive therapy and nonetheless have cure rates below 50%. Treatment with the ch14.18 monoclonal antibody (dinutuximab beta), directed against the GD2 disialoganglioside, improved 5-year event-free survival in high-risk patients when administered in postconsolidation therapy and was recently implemented in standard therapy. Relapse still occurred in 57% of these patients, necessitating new therapeutic options. Bispecific trifunctional antibodies (trAbs) are IgG-like molecules directed against T cells and cancer surface antigens, redirecting T cells (via their CD3 specificity) and accessory immune cells (via their functioning Fc-fragment) toward tumor cells. We sought proof-of-concept for GD2/CD3-directed trAb efficacy against neuroblastoma. Methods: We used two GD2-specific trAbs differing only in their CD3-binding specificity: EKTOMUN (GD2/human CD3) and SUREK (GD2/mouse Cd3). This allowed trAb evaluation in human and murine experimental settings. Tumor-blind trAb and the ch14.18 antibody were used as controls. A coculture model of human peripheral blood mononuclear cells (PBMCs) and neuroblastoma cell lines was established to evaluate trAb antitumor efficacy by assessing expression of T-cell surface markers for activation, proinflammatory cytokine release and cytotoxicity assays. Characteristics of tumor-infiltrating T cells and response of neuroblastoma metastases to SUREK treatment were investigated in a syngeneic immunocompetent neuroblastoma mouse model mimicking minimal residual disease. Results: We show that EKTOMUN treatment caused effector cell activation and release of proinflammatory cytokines in coculture with neuroblastoma cell lines. Furthermore, EKTOMUN mediated GD2-dependent cytotoxic effects in human neuroblastoma cell lines in coculture with PBMCs, irrespective of the level of target antigen expression. This effect was dependent on the presence of accessory immune cells. Treatment with SUREK reduced the intratumor Cd4/Cd8 ratio and activated tumor infiltrating T cells in vivo. In a minimal residual disease model for neuroblastoma, we demonstrated that single-agent treatment with SUREK strongly reduced or eliminated neuroblastoma metastases in vivo. SUREK as well as EKTOMUN demonstrated superior tumor control compared with the anti-GD2 antibody, ch14.18. Conclusions: Here we provide proof-of-concept for EKTOMUN preclinical efficacy against neuroblastoma, presenting this bispecific trAb as a promising new agent to fight neuroblastoma

A longitudinal study of risk factors for the occurrence, duration and severity of menstrual cramps in a cohort of college women

To describe how menstrual cramps vary from cycle to cycle within a woman over time. To examine the influence of weight and lifestyle factors on occurrence, duration, and severity of menstrual pain. Design A one-year prospective menstrual diary study. Participants One hundred and sixty-five women aged 17 to 19 years entering a local university in 1985. Main outcome measures The occurrence, length, and maximum severity of pain during a menstrual period. Results Menstrual pain occurred during 71.6% of observed menstrual bleeds, most commonly beginning the first day of menses. The median duration was two days. Sixty percent of women reported at least one episode of severe pain, while 13% reported severe pain more than half the time. Earlier age at menarche and long menstrual periods increased the occurrence, duration and severity of pain. In smokers, cramps tended to last longer. Being overweight was an important risk factor for menstrual cramps and doubled the odds of having a long pain episode. Frequent alcohol consumption decreased the probability of having menstrual cramps, but in women who had pain it increased duration and severity. Physical activity was not associated with any pain parameter. Conclusions Women who have pain lasting three days are an important target group for prophylactic therapy. The occurrence and severity of menstrual cramps is influenced by potentially modifiable characteristics including weight, smoking, and alcohol consumption. Doctors may wish to counsel women presenting with dysmenorrhoea about the importance of healthy lifestyles and about the inefficacy of alcohol consumption as a treatment for dysmenorrhoea.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73220/1/j.1471-0528.1996.tb09597.x.pd

Identification of 14-3-3γ as a Mieap-interacting protein and its role in mitochondrial quality control

Mieap, a p53-inducible protein, controls mitochondrial integrity by inducing the accumulation of lysosomal proteins within mitochondria. This phenomenon is designated MALM, for Mieap-induced accumulation of lysosome-like organelles within mitochondria. To identify this novel Mieap-interacting protein(s), we performed a two-dimensional image-converted analysis of liquid chromatography and mass spectrometry (2DICAL) on the proteins immunoprecipitated by an anti-Mieap antibody. We indentified 14-3-3γ as one of the proteins that was included in the Mieap-binding protein complex when MALM was induced. The interaction between Mieap and 14-3-3γ was confirmed on the exogenous and endogenous proteins. Interestingly, 14-3-3γ was localized within mitochondria when MALM occurred. A 14-3-3γ deficiency did not affect the accumulation of Mieap and lysosomal proteins within mitochondria, but dramatically inhibited the elimination of oxidized mitochondrial proteins. These results suggest that 14-3-3γ plays a critical role in eliminating oxidized mitochondrial proteins during the MALM process by interacting with Mieap within mitochondria

Biobehavioral research on nicotine use in women

More American women are taking up smoking than men and fewer are quitting; if current trends continue, rates for women will surpass those for men by the mid-1990's. But ironically, much of what is known about the biobehavioural aspects of smoking is based on research using male subjects. The present paper reviews evidence suggesting that: (1) women may differ from men with regard to nicotine intake and/or effects; (2) nicotine intake and effects may be influenced by menstrual cycle phase; (3) oral contraceptive use and estrogen replacement therapy may affect intake and effects of nicotine; (4) the effects of chronic nicotine use on female reproductive endocrinology may have implications for the reinforcement of smoking; and (5) pharmacological agents used to treat smoking may have different effects in women than in men. Guidelines and suggestions are presented by future biobehavioural research in women, including standardization of assessment procedures, attention to the use of appropriate controls, and use of pharmacological probes.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73978/1/j.1360-0443.1991.tb01802.x.pd

Symptom patterns in women with premenstrual syndrome complaints: a prospective assessment using a marker for ovulation and screening criteria for adequate ovarian function

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72929/1/j.1365-2648.1991.tb01727.x.pd

Coenzyme A-transferase-independent butyrate re-assimilation in Clostridium acetobutylicum - evidence from a mathematical model

The hetero-dimeric CoA-transferase CtfA/B is believed to be crucial for the metabolic transition from acidogenesis to solventogenesis in Clostridium acetobutylicum as part of the industrial-relevant acetone-butanol-ethanol (ABE) fermentation. Here, the enzyme is assumed to mediate re-assimilation of acetate and butyrate during a pH-induced metabolic shift and to faciliate the first step of acetone formation from acetoacetyl-CoA. However, recent investigations using phosphate-limited continuous cultures have questioned this common dogma. To address the emerging experimental discrepancies, we investigated the mutant strain Cac-ctfA398s::CT using chemostat cultures. As a consequence of this mutation, the cells are unable to express functional ctfA and are thus lacking CoA-transferase activity. A mathematical model of the pH-induced metabolic shift, which was recently developed for the wild type, is used to analyse the observed behaviour of the mutant strain with a focus on re-assimilation activities for the two produced acids. Our theoretical analysis reveals that the ctfA mutant still re-assimilates butyrate, but not acetate. Based upon this finding, we conclude that C. acetobutylicum possesses a CoA-tranferase-independent butyrate uptake mechanism that is activated by decreasing pH levels. Furthermore, we observe that butanol formation is not inhibited under our experimental conditions, as suggested by previous batch culture experiments. In concordance with recent batch experiments, acetone formation is abolished in chemostat cultures using the ctfa mutant

WHO systematic review of prevalence of chronic pelvic pain: a neglected reproductive health morbidity

BACKGROUND: Health care planning for chronic pelvic pain (CPP), an important cause of morbidity amongst women is hampered due to lack of clear collated summaries of its basic epidemiological data. We systematically reviewed worldwide literature on the prevalence of different types of CPP to assess the geographical distribution of data, and to explore sources of variation in its estimates. METHODS: We identified data available from Medline (1966 to 2004), Embase (1980 to 2004), PsycINFO (1887 to 2003), LILACS (1982 to 2004), Science Citation index, CINAHL (January 1980 to 2004) and hand searching of reference lists. Two reviewers extracted data independently, using a piloted form, on participants' characteristics, study quality and rates of CPP. We considered a study to be of high quality (valid) if had at least three of the following features: prospective design, validated measurement tool, adequate sampling method, sample size estimation and response rate >80%. We performed both univariate and multivariate meta-regression analysis to explore heterogeneity of results across studies. RESULTS: There were 178 studies (459975 participants) in 148 articles. Of these, 106 studies were (124259 participants) on dysmenorrhoea, 54 (35973 participants) on dyspareunia and 18 (301756 participants) on noncyclical pain. There were only 19/95 (20%) less developed and 1/45 (2.2%) least developed countries with relevant data in contrast to 22/43 (51.2%) developed countries. Meta-regression analysis showed that rates of pain varied according to study quality features. There were 40 (22.5%) high quality studies with representative samples. Amongst them, the rate of dysmenorrhoea was 16.8 to 81%, that of dyspareunia was 8 to 21.8%, and that for noncyclical pain was 2.1 to 24%. CONCLUSION: There were few valid population based estimates of disease burden due to CPP from less developed countries. The variation in rates of CPP worldwide was due to variable study quality. Where valid data were available, a high disease burden of all types of pelvic pain was found