Truncation of annexin A1 is a regulatory lever for linking epidermal growth factor signaling with cytosolic phospholipase A2 in normal and malignant squamous epithelial cells

Regulation of cell growth and apoptosis is one of the pleiotropic functions of annexin A1 (ANXA1). Although previous reports on the overexpression of ANXA1 in many human cancers and on growth suppression and/or induction of apoptosis by ANXA1 may indicate the tumor-suppressive nature of ANXA1, molecular mechanisms of the function of ANXA1 remain largely unknown. Here we provide evidence that ANXA1 mechanistically links the epidermal growth factor-triggered growth signal pathway with cytosolic phospholipase A(2) (cPLA(2)), an initiator enzyme of the arachidonic acid cascade, through interaction with S100A11 in normal human keratinocytes (NHK). Ca(2+)-dependent binding of S100A11 to ANXA1 facilitated the binding of the latter to cPLA(2), resulting in inhibition of cPLA(2) activity, which is essential for the growth of NHK. On exposure of NHK to epidermal growth factor, ANXA1 was cleaved solely at Trp(12), and this cleavage was executed by cathepsin D. In squamous cancer cells, this pathway was shown to be constitutively activated. The newly found mechanistic intersection may be a promising target for establishing new measures against human cancer and other cell growth disorders.     

Affecting Pseudomonas aeruginosa Phenotypic Plasticity by Quorum Sensing Dysregulation Hampers Pathogenicity in Murine Chronic Lung Infection

In Pseudomonas aeruginosa quorum sensing (QS) activates the production of virulence factors, playing a critical role in pathogenesis. Multiple negative regulators modulate the timing and the extent of the QS response either in the pre-quorum or post-quorum phases of growth. This regulation likely increases P. aeruginosa phenotypic plasticity and population fitness, facilitating colonization of challenging environments such as higher organisms. Accordingly, in addition to the factors required for QS signals synthesis and response, also QS regulators have been proposed as targets for anti-virulence therapies. However, while it is known that P. aeruginosa mutants impaired in QS are attenuated in their pathogenic potential, the effect of mutations causing a dysregulated timing and/or magnitude of the QS response has been poorly investigated so far in animal models of infection. In order to investigate the impact of QS dysregulation on P. aeruginosa pathogenesis in a murine model of lung infection, the QteE and RsaL proteins have been selected as representatives of negative regulators controlling P. aeruginosa QS in the pre- and post-quorum periods, respectively. Results showed that the qteE mutation does not affect P. aeruginosa lethality and ability to establish chronic infection in mice, despite causing a premature QS response and enhanced virulence factors production in test tube cultures compared to the wild type. Conversely, the post-quorum dysregulation caused by the rsaL mutation hampers the establishment of P. aeruginosa chronic lung infection in mice without affecting the mortality rate. On the whole, this study contributes to a better understanding of the impact of QS regulation on P. aeruginosa phenotypic plasticity during the infection process. Possible fallouts of these findings in the anti-virulence therapy field are also discussed.     

Publication of clinical trials supporting successful new drug applications: a literature analysis

Background

The United States (US) Food and Drug Administration (FDA) approves new drugs based on sponsor-submitted clinical trials. The publication status of these trials in the medical literature and factors associated with publication have not been evaluated. We sought to determine the proportion of trials submitted to the FDA in support of newly approved drugs that are published in biomedical journals that a typical clinician, consumer, or policy maker living in the US would reasonably search.

Methods and Findings

We conducted a cohort study of trials supporting new drugs approved between 1998 and 2000, as described in FDA medical and statistical review documents and the FDA approved drug label. We determined publication status and time from approval to full publication in the medical literature at 2 and 5 y by searching PubMed and other databases through 01 August 2006. We then evaluated trial characteristics associated with publication. We identified 909 trials supporting 90 approved drugs in the FDA reviews, of which 43% (394/909) were published. Among the subset of trials described in the FDA-approved drug label and classified as “pivotal trials” for our analysis, 76% (257/340) were published. In multivariable logistic regression for all trials 5 y postapproval, likelihood of publication correlated with statistically significant results (odds ratio [OR] 3.03, 95% confidence interval [CI] 1.78–5.17); larger sample sizes (OR 1.33 per 2-fold increase in sample size, 95% CI 1.17–1.52); and pivotal status (OR 5.31, 95% CI 3.30–8.55). In multivariable logistic regression for only the pivotal trials 5 y postapproval, likelihood of publication correlated with statistically significant results (OR 2.96, 95% CI 1.24–7.06) and larger sample sizes (OR 1.47 per 2-fold increase in sample size, 95% CI 1.15–1.88). Statistically significant results and larger sample sizes were also predictive of publication at 2 y postapproval and in multivariable Cox proportional models for all trials and the subset of pivotal trials.

Conclusions

Over half of all supporting trials for FDA-approved drugs remained unpublished ≥ 5 y after approval. Pivotal trials and trials with statistically significant results and larger sample sizes are more likely to be published. Selective reporting of trial results exists for commonly marketed drugs. Our data provide a baseline for evaluating publication bias as the new FDA Amendments Act comes into force mandating basic results reporting of clinical trials.     

Chromosome analysis by spectral karyotyping of spermatozoa from an oligoasthenozoospermic carrier of a 10; 21 reciprocal translocation

Cytogenetic analysis of germ-line cells prior to intracytoplasmic sperm injection (ICSI) treatment is thought to be necessary for infertile males with an identified chromosomal abnormality. We analyzed the chromosomal karyotype of human spermatozoa from an oligoasthenozoospermic carrier of a reciprocal translocation t(10; 21). Cytogenetic analysis of 39 spermatozoa was performed by spectral karyotyping (SKY) and by ICSI into mouse oocytes. The motile morphologically normal spermatozoa were injected into mouse oocytes. Of these spermatozoa, 38 (97.4%) were activated. Twenty-one (53.8%) of the activated oocytes formed two pronuclei. Metaphase chromosome spreads from 13 spermatozoa were analyzed. Only one spermatozoon was normal and 2 spermatozoa exhibited balanced translocation. Nine and one spermatozoa showed abnormalities related and unrelated to the translocation, respectively. The numbers of normal/balanced spermatozoa were lower than those in previous reports analyzing reciprocal translocations using a previously described technique involving penetrated golden hamster oocytes. After genetic counseling with the carrier and his partner, ICSI treatment was performed. Healthy female and male infants were delivered at 37 weeks gestation via a Caesarean section. The female infant was a carrier of the reciprocal translocation and the male infant was confirmed normal on prenatal diagnosis at 16 weeks gestation. For genetic counseling prior to ICSI treatment, the incidence of unbalanced type spermatozoa after swim-up or Percoll gradient treatment should be investigated and discussed with couples having fertility problems related to oligozoospermia autosomal structural abnormalities.     

Genetic analysis of the yeastCandida maltosa by means of induced parasexual processes

The usefulness of hybridization by protoplast fusion and mitotic segregation for the genetic analysis of the imperfect fodder yeastCandida maltosa was tested. Mitotically stable fusion hybrids were obtained with frequencies between 10^–6 and 10^–7. Complementation tests were performed by protoplast fusion. Substances that are known to induce frequent mitotic segregation in other yeast species such as benomyl, p-fluorophenylalanine, and acriflavine were ineffective inC. maltosa. UV irradiation induced mitotic segregation in up to 10%. This agent induced mainly mitotic crossing over inC. maltosa. Our data enabled the construction of the linkage group I with the sequenceCEN-ade-26-pro-1.