The antibody-mediated targeted delivery of interleukin-12 (IL12) to the EDB domain of fibronectin, a marker of angiogenesis, is a promising avenue for enhancing the therapeutic index of this anticancer cytokine. Previous experiments, based on sequential fusion of a single-chain IL12 derivative to the anti-EDB antibody fragment scFv(L19) had yielded a therapeutic fusion protein [IL12-scFv(L19)-FLAG], which displayed an impressive therapeutic activity in murine models of cancer, in spite of a tumor uptake, which was less efficient compared to the parental unmodified scFv(L19). In this article, we describe the comparative analysis of 3 recombinant fusion proteins comprising the scFv(L19) and IL12 moieties. One of them, in which the p40 and p35 form a covalent heterodimer and in which each subunit is fused to a molecule of scFv(L19), displays an excellent tumor targeting performance in vivo, as assessed by quantitative biodistribution analysis, and a potent anti-tumor effect, superior to the one of IL12-scFv(L19)-FLAG. These results may have a clinical impact, considering the fact that the tumor targeting ability of scFv(L19) in patients with cancer has been demonstrated using scintigraphic methods, and that 2 scFv(L19)-based antibody-cytokine fusion are currently entering clinical trials. The targeted delivery of cytokines to the tumor environment represents a promising approach for enhancing the therapeutic index of these biopharmaceuticals, which are often hampered by severe toxicity even at low doses. [1][2][3] In the most common implementation, ligands (such as peptides, antibody fragments or full antibodies) specific to tumor-associated antigens are genetically fused to cytokines in order to mediate a preferential accumulation of the fusion protein at the tumor site 1,2,4-8 ; For these applications, tumor vascular antigens may represent a particularly attractive class of targets, because of their ready accessibility for biopharmaceuticals which are administered intravenously. 9 In the past, we have generated in collaboration with the group of Luciano Zardi a high-affinity human monoclonal antibody 10 specific to the EDB domain of fibronectin, a marker of angiogenesis. 11-14 The L19 antibody was shown to selectively localize to tumor blood vessels in animal models of cancer [15][16][17][18] and in patients with aggressive solid tumors. 19 The L19 antibody in single-chain Fv recombinant antibody format (''scFv(L19)'') was successfully fused to several cytokines 20 and pro-coagulant factors, 21 or chemically conjugated to radionuclides, 17,18 drugs [D.N., unpublished], photosensitizers 22,23 and enzymes. 24 Three of these derivatives (the L19 antibody in SIP (Small Immuno-Protein) format 18 labeled with 131 I, scFv(L19) fused to IL2 and scFv(L19) fused to TNF-a) are currently in clinical development.The heterodimeric cytokine interleukin-12 (IL-12) is a key mediator of innate and cellular immunity with potent antitumor and antimetastatic activity, [25][26][27] and is currently being tested in Phase II clinical tr...

IntroductionPseudomonas aeruginosa frequently causes nosocomial pneumonia and is associated with poor outcome. The purpose of this study was to assess the prevalence and clinical outcome of nosocomial pneumonia caused by serotype-specific P. aeruginosa in critically ill patients under appropriate antimicrobial therapy management.MethodsA retrospective, non-interventional epidemiological multicenter cohort study involving 143 patients with confirmed nosocomial pneumonia caused by P. aeruginosa. Patients were analyzed for a period of 30 days from time of nosocomial pneumonia onset. Fourteen patients fulfilling the same criteria from a phase IIa studyconducted at the same time/centers were included in the prevalence calculations but not in the clinical outcome analysis.ResultsThe prevalence of serotypes was: O6 (29%), O11 (23%), O10 (10%), O2 (9%), and O1 (8%). Serotypes with a prevalence of less than 5% were found in 13% of patients, 8% were classified as not typeable. Across all serotypes, 19% mortality, 70% clinical resolution, 11% clinical continuation, and 5% clinical recurrence were recorded. Age and higher APACHE II (Acute Physiology and Chronic Health Evaluation II) were predictive risk factors associated with probability of death and lower clinical resolution for P. aeruginosa nosocomial pneumonia. Mortality tends to be higher with O1 (40%) and lower with O2 (0%); clinical resolution tends to be better with O2 (82%) compared to other serotypes. Persisting pneumonia with O6 and O11 was, respectively, 8% and 21%; clinical resolution with O6 and O11 was, respectively, 75% and 57%.ConclusionsIn P. aeruginosa nosocomial pneumonia, the most prevalent serotypes were O6 and O11. Further studies including larger group sizes are needed to correlate clinical outcome with virulence factors of P. aeruginosa in patients with nosocomial pneumonia caused by various serotypes; and to compare O6 and O11, the two serotypes most frequently encountered in critically ill patients.

Functional thrombin receptors are present on endothelium and SMCs of IMA and SV. Endothelial thrombin receptors mediate relaxation in IMA but not SV. Thrombin causes much more pronounced contraction and proliferation in SMCs of SV than IMA independent of tethered receptors, suggesting other thrombin receptors exist. These differences of thrombin receptor activation in IMA and SV may be important in the development of and therapy for graft disease.

These data suggest that panobacumab is safe, with a pharmacokinetic profile similar to that in healthy volunteers. It was associated with high clinical cure and survival rates in patients developing nosocomial P. aeruginosa O11 pneumonia. We concluded that these promising results warrant further trials.

Pseudomonas aeruginosa is a common cause of nosocomial infections and is associated with high rates of mortality. In order to facilitate rapid and sensitive identification of the most prevalent serotypes of P. aeruginosa, we have developed a 4-valent real-time PCR-based assay using oligonucleotides specific for open-reading frames constituting the O-antigen-specific lipopolysaccharide loci of P. aeruginosa. The assay simultaneously detects and differentiates between each of the four serotypes IATS-O1, -O6, -O11 and serogroup 2 (IATS-O2, -O5, and -O16) with high sensitivity and specificity in a single-tube reaction. No cross-reactivity was observed with other serotypes of P. aeruginosa or other microbial species, and reproducibility was demonstrated regardless of template, i.e. purified DNA, bacterial culture and clinical specimens (broncho-alveolar lavage). The limit of detection of the assay was approximately 100 copies per reaction for IATS-O1, -O2 and -O11, and 50 copies per reaction for IATS-O6. Comparison of the assay specificity with a commercially available slide agglutination kit showed consistent results; however, the number of non-typable isolates was reduced by 15 % using the genotyping assay. Use of the 4-valent genotyping assay in the context of a clinical trial resulted in identification of pneumonia patients positive for the IATS-O11 serotype, and hence eligible for therapy with panobacumab (an investigational monoclonal antibody against the O-polysaccharide of serotype IATS-O11).