Drug Levels in the Maternal Serum, Cord Blood and Breast Milk of a Ustekinumab-Treated Patient with Crohn's Disease.

Ustekinumab [UST] therapy during pregnancy has not yet been extensively evaluated in patients with Crohn's disease. Here, we present the case of a 24-year-old woman with therapy-refractory Crohn's disease, who was treated with UST until Week 30 of pregnancy and successfully delivered a healthy baby boy, who had normal development in the follow-up period of one year. The cord blood UST level was markedly higher than the measured maternal serum drug level. The trough level in the breast milk after re-initiating postpartum UST therapy was initially in the same range as the corresponding serum trough level, and then decreased during maintenance therapy. This is one of the first reports describing the drug levels in the breast milk after re-initiating UST treatment in a Crohn's disease patient.

Proteomic pattern of implantative human endometrial fluid in in vitro fertilization cycles.

PURPOSE: To assess whether there are proteins in endometrial fluid aspirate (EFA) that predict implantation. METHODS: The population under study consisted of 285 women undergoing embryo transfer (ET). Endometrial fluid aspiration was performed immediately before ET. Results of proteomic analysis of EFA were compared between 33 cases who achieved pregnancy and 33 who did not. Samples were analysed by 2D electrophoresis and mass spectrometry. Blood samples were studied by ELISA Pregnancy rates and maternal complications were compared to those in women refusing aspiration. RESULTS: We found 23 proteins differentially expressed in the EFA in conception cycles: 4 up-regulated proteins and 19 down-regulated (FC = 0.31 0.78) (among others, arginase-1, actin B, PARK-7, cofilin-1, stathmin, annexin-2 and CAPZB). Among the five studied proteins that were differentially expressed in EFA, none was differentially expressed in serum. The aspiration procedure had no impact on pregnancy rate. No maternal complications were reported. CONCLUSIONS: We found a very different protein profile in implantative cycles, the majority of proteins being down-regulated. This probably reflects a different endometrial functional status, more favourable to implantation. EFA proteomic analysis could be a useful tool in the planning ET strategies.

A new biomarker panel in bronchoalveolar lavage for an improved lung cancer diagnosis.

INTRODUCTION: The enormous biological complexity and high mortality rate of lung cancer highlights the need for new global approaches for the discovery of reliable early diagnostic biomarkers. The study of bronchoalveolar lavage samples by proteomic techniques could identify new lung cancer biomarkers and may provide promising noninvasive diagnostic tools able to enhance the sensitivity of current methods. METHODS: First, an observational prospective study was designed to assess protein expression differences in bronchoalveolar lavages from patients with (n = 139) and without (n = 49) lung cancer, using two-dimensional gel electrophoresis and subsequent protein identification by mass spectrometry. Second, validation of candidate biomarkers was performed by bead-based immunoassays with a different patient cohort (204 patients, 48 controls). RESULTS: Thirty-two differentially expressed proteins were identified in bronchoalveolar lavages, 10 of which were confirmed by immunoassays. The expression levels of APOA1, CO4A, CRP, GSTP1, and SAMP led to a lung cancer diagnostic panel that reached 95% sensitivity and 81% specificity, and the quantification of STMN1 and GSTP1 proteins allowed the two main lung cancer subtypes to be discriminated with 90% sensitivity and 57% specificity. CONCLUSIONS: Bronchoalveolar lavage represents a promising noninvasive source of lung cancer specific protein biomarkers with high diagnostic accuracy. Measurement of APOA1, CO4A, CRP, GSTP1, SAMP, and STMN1 in this fluid may be a useful tool for lung cancer diagnosis, although a further validation in a larger clinical set is required for early stages.

Influence of immunogenicity on the efficacy of longterm treatment of spondyloarthritis with infliximab.

BACKGROUND: Infliximab (IFX) is a monoclonal antibody against tumour necrosis factor α that is effective for treating spondyloarthritis (SpA). However, after initial success of the drug some patients lose responsiveness or develop infusion reactions, which may be related to the development of antibodies against the drug. OBJECTIVE: To investigate the clinical relevance of antibodies to infliximab (ATI) formation in patients with SpA undergoing IFX treatment over a prolonged period. METHODS: 94 patients with SpA treated with IFX from 1999 to 2010 were studied. Their clinical characteristics, serum trough IFX levels and ATI status were evaluated for a mean of 6.99 (95% CI:6.28 to 7.7) years. Clinical activity and improvement were measured using the Ankylosing Spondylitis Disease Activity Score (ASDAS): inactive <1.3, moderate ≥1.3 and <2.1, high ≥2.1-≤3.5, and very high >3.5 at three time points (6 months, 12 months and >4 years). RESULTS: ATI were detected in 24 (25.5%) patients. The patients with ATI had higher ASDAS scores than those without ATI (2.55±0.89 vs 1.79±1.04, p=0.038 at 6 months; 1.95±0.67 vs 1.67±0.71, p=0.042 at 1 year; 2.52±0.99 vs 1.53±0.81, p=0.024 at >4 years). Eleven patients (12%) developed infusion-related reactions, and of these, ATI were present in eight patients (73%). The patients with infusion-related reactions had higher ATI titres (median 12 931 AU/ml, IQR 853-82 437) vs median 2454 AU/ml, IQR 449-7718, p=0.028) and shorter survival (4.25 years vs 8.19 years, p<0.001). ATI development occurred more frequently in the patients not receiving methotrexate (20/58 (34.5%) vs 4/36 (11.1%), p=0.011). CONCLUSION: In patients with SpA treated with IFX, ATI formation is associated with a poor clinical response, the appearance of infusion reactions and the discontinuation of treatment.

Comprehensive proteomic analysis of human endometrial fluid aspirate.

The endometrial fluid is a noninvasive sample which contains numerous secreted proteins representative of endometrial function and reflects the state of the endometrium. In this study, we describe, for the first time, a comprehensive catalogue of proteins of the endometrial fluid during the secretory phase of the menstrual cycle. To achieve this objective, three different but complementary strategies were used: First, in-solution digestion followed by reverse phase high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS); second, protein separation by denaturing one-dimensional electrophoresis (SDS-PAGE) followed by HPLC-MS/MS analysis. Finally, two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) followed by MALDI-TOF/TOF analysis. The combination of the three strategies led to the successful identification of 803 different proteins in the International Protein Index (IPI) human database (v3.48). An extensive description of the endometrial fluid proteome will help provide the basis for a better understanding of a number of diseases and processes, including endometriosis, endometrial cancer and embryo implantation. We believe that the thorough catalogue of proteins presented here can serve as a valuable reference for the study of embryo implantation and for future biomarker discovery involved in pathologic alterations of endometrial function.

Characterization of the N-terminal domain of NrtC, the ATP-binding subunit of ABC-type nitrate transporter of the cyanobacterium Phormidium laminosum.

The N-terminal domain of NrtC, the ATP-binding subunit of nitrate/nitrite ABC-transporter in the cyanobacterium Phormidium laminosum, has been expressed in Escherichia coli as a histidine-tagged fusion protein (His(6)NrtC1). Binding of ATP to the pure His(6)NrtC1 was characterized using the nucleotide analogue TNP-ATP [2'(3')-O-(2,4,6-trinitrophenyl) adenosine 5'-triphosphate]. Fluorescence assays showed that His(6)NrtC1 specifically binds Mg(2+) TNP-ATP with high affinity, binding being dependent on protein concentration. The presence of ATP prevents the covalent modification of His(6)NrtC1 by fluorescein 5'-isothiocyanate (FITC), suggesting that this probe reacts at the nucleotide-binding site of NrtC. The active form of the truncated NrtC is a dimer that shows high affinity for TNP-ATP (K(d)=0.76+/-0.1 microM). Evidence for the presence of two nucleotide-binding sites per dimer protein is given. Our results indicate that nucleotide binding is strongly dependent on the dimerization of NrtC and that the N-terminal domain of the protein contains the binding site for ATP. No ATPase activity catalyzed in vitro by the truncated subunit was detected.