A directed genome evolution method to enhance hydrogen production in .

Nitrogenase-dependent H production by photosynthetic bacteria, such as , has been extensively investigated. An important limitation to increase H production using genetic manipulation is the scarcity of high-throughput screening methods to detect possible overproducing mutants. Previously, we engineered strains that emitted fluorescence in response to H and used them to identify mutations in the nitrogenase Fe protein leading to H overproduction.

Genetic and Biochemical Analysis of the Molybdenum Storage Protein.

The N fixing bacterium carries a molybdenum storage protein, referred to as MoSto, able to bind 25-fold more Mo than needed for maximum activity of its Mo nitrogenase. Here we have investigated a plausible role of MoSto as obligate intermediate in the pathway that provides Mo for the biosynthesis of nitrogenase iron-molybdenum cofactor (FeMo-co). The FeMo-co synthesis and insertion assay demonstrated that purified MoSto functions as Mo donor and that direct interaction with FeMo-co biosynthetic proteins stimulated Mo donation.

Expression of a functional oxygen-labile nitrogenase component in the mitochondrial matrix of aerobically grown yeast.

The extreme sensitivity of nitrogenase towards oxygen stands as a major barrier to engineer biological nitrogen fixation into cereal crops by direct nif gene transfer. Here, we use yeast as a model of eukaryotic cell and show that aerobically grown cells express active nitrogenase Fe protein when the NifH polypeptide is targeted to the mitochondrial matrix together with the NifM maturase. Co-expression of NifH and NifM with Nif-specific Fe-S cluster biosynthetic proteins NifU and NifS is not required for Fe protein activity, demonstrating NifH ability to incorporate endogenous mitochondrial Fe-S clusters.

Randomized phase II study comparing gemcitabine plus dacarbazine versus dacarbazine alone in patients with previously treated soft tissue sarcoma: a Spanish Group for Research on Sarcomas study.

Purpose: To assess the activity and toxicity of the combination of gemcitabine plus dacarbazine (DTIC) in patients with advanced soft tissue sarcoma (STS) in a randomized, multicenter, phase II study using DTIC alone as a control arm.

Patients And Methods: Patients with previously treated advanced STS were randomly assigned to receive either fixed-dose rate gemcitabine (10 mg/m2/min) at 1800 mg/m2 followed by DTIC at 500 mg/m2 every 2 weeks, or DTIC alone at 1200 mg/m2 every 3 weeks. The primary end point of the study was progression-free rate (PFR) at 3 months.

Trabectedin in pre-treated patients with advanced or metastatic soft tissue sarcoma: a phase II study evaluating co-treatment with dexamethasone.

Purpose: This study assesses the efficacy, toxicity and pharmacokinetic profile of trabectedin with or without prophylactic dexamethasone co-treatment in patients with recurrent advanced soft tissue sarcoma (STS).

Patients And Methods: Patients were randomized to receive trabectedin as a 3-h infusion every 3 weeks with dexamethasone or placebo in the first cycle, with the alternate in the second cycle and with the patient's choice subsequently. Due to toxicity, the randomized design was modified to open-label to make dexamethasone mandatory and the initial dose (1,650 μg/m(2)) was reduced to 1,500 μg/m(2) and then to 1,300 μg/m(2).

Efficacy of sequential high-dose doxorubicin and ifosfamide compared with standard-dose doxorubicin in patients with advanced soft tissue sarcoma: an open-label randomized phase II study of the Spanish group for research on sarcomas.

Purpose: To assess the progression-free survival (PFS) and antitumor response to standard-dose doxorubicin compared with sequential dose-dense doxorubicin and ifosfamide in first-line treatment of advanced soft tissue sarcoma.

Patients And Methods: Patients with measurable advanced soft tissue sarcoma, Eastern Cooperative Oncology Group (ECOG) performance status (PS) < 2, between the ages 18 and 65 years, and with adequate bone marrow, liver, and renal function were entered in the study. The stratifications were: ECOG PS (0 v 1), location of metastases, and potentially resectable disease.

Pulmonary toxicity in patients treated with gemcitabine plus vinorelbine or docetaxel for advanced non-small cell lung cancer: outcome data on a randomized phase II study.

Studies with the gemcitabine/vinorelbine (GV) or the gemcitabine/docetaxel (GD) combinations have shown similar efficacy and less toxicity compared to platinum-based chemotherapies, in patients with advanced non-small-cell lung cancer (NSCLC). The present trial was designed to test the efficacy and safety of both, GV and GD, combinations. Chemotherapy-naïve patients (n=39)or=60% and adequate hematological, renal and hepatic function were randomly assigned to receive G 1,000 mg/m2+either V 25 mg/m2 or D 35 mg/m2 (all of which were administered i.

Cisplatin plus gemcitabine with or without vinorelbine as induction chemotherapy prior to radical locoregional treatment for patients with stage III non-small-cell lung cancer (NSCLC): results of a prospective randomized study.

To evaluate possible improvement in objective response of adding vinorelbine (V) to the combination of cisplatin/gemcitabine (CG) in induction chemotherapy for stage III NSCLC, patients (n=154) aged < or =75 years, Karnofsky index > or =70%, were stratified by stage (IIIA versus IIIB) and randomly assigned to receive: C (50mg/m(2) i.v.) plus G (1250mg/m(2) i.

Phase II study with the combination of gemcitabine and DTIC in patients with advanced soft tissue sarcomas.

Purpose: Based on the promising results of a Phase I study with a combination of gemcitabine and DTIC performed in advanced soft tissue sarcoma (ASTS) patients, and due to the limited efficacy of second or third line therapies in those patients, we designed a Phase II study to determine the activity of this new regimen.

Methods: Patients with ASTS, measurable disease, pretreated with chemotherapy, received gemcitabine 1,800 mg/m2 infused over 180 min followed by DTIC 500 mg/m2 (one cycle), every 2 weeks. The pharmacokinetics (PK) of gemcitabine and 2',2'-difluorodeoxyuridine (dFdU), and the accumulation of gemcitabine triphosphate (dFdCTP) by peripheral blood mononuclear cells were studied.

Deletions affecting codons 557-558 of the c-KIT gene indicate a poor prognosis in patients with completely resected gastrointestinal stromal tumors: a study by the Spanish Group for Sarcoma Research (GEIS).

Purpose: To explore the prognostic value of mutations in c-KIT and PDGFR-alpha genes with respect to relapse-free survival (RFS) in patients with gastrointestinal stromal tumors (GIST). We have investigated the prognostic relevance of the type and position of the mutations, in addition to other clinicopathologic factors, in a large series of patients with GIST.

Methods: For this study, 162 patients were selected according to the following criteria: completely resected tumors with negative margins attended between 1994 and 2001; no metastasis at diagnosis; tumor larger than 2 cm, c-KIT-positive immunostaining; and no other primary tumors.

A phase II trial of temozolomide as a 6-week, continuous, oral schedule in patients with advanced soft tissue sarcoma: a study by the Spanish Group for Research on Sarcomas.

Background: The objective of this study was to evaluate the activity and toxicity of temozolomide given as an extended schedule in patients with advanced sarcoma.

Methods: Forty-nine patients with pretreated soft tissue sarcoma (the STS arm) and 18 patients with previously untreated gastrointestinal stromal tumor (the GIST arm) were enrolled onto a 2-arm, multicenter, Phase II study between November 1999 and July 2001. Temozolomide was administered on a 6-week, continuous, oral schedule at a dose of 75 mg/m2 per day in 41 patients and, after an amendment, at a dose of 100 mg/m2 per day in 22 patients.

Gemcitabine and vinorelbine (GV) versus cisplatin, gemcitabine and vinorelbine (CGV) as first-line treatment in advanced non small cell lung cancer: results of a prospective randomized phase II study.

The objective of this study was to assess whether adding cisplatin to gemcitabine/vinorelbine combination improves the clinical outcome in patients with non-small-cell lung cancer (NSCLC). Chemotherapy-naïve patients with advanced NSCLC; age < or = 75 years: Karnofsky performance status > or = 60%, and with adequate hematological, renal and hepatic function, were randomized into 2 treatment groups to receive Gemcitabine 1250 mg/m2 + vinorelbine 30 mg/m2 (GV group), or cisplatin 50 mg/m2 + gemcitabine 1000 mg/m2 + vinorelbine 25 mg/m2 (CGV group). All drugs were administered on days 1 and 8 every three weeks: From September 1999 to March 2003, 114 patients were enrolled.

Phase I clinical trial of fixed-dose rate infusional gemcitabine and dacarbazine in the treatment of advanced soft tissue sarcoma, with assessment of gemcitabine triphosphate accumulation.

Background: In the current study, the authors set out to determine the dose-limiting toxicity (DLT) and maximum tolerated dose (MTD) associated with a combination of gemcitabine and dacarbazine (DTIC) in patients with advanced soft tissue sarcoma (ASTS), to obtain preliminary information on the activity of this combination, and to explore possible pharmacodynamic interactions between gemcitabine and DTIC.

Methods: Every 2 weeks, 22 patients with refractory ASTS received fixed-dose rate gemcitabine (10 mg/m2/min) at escalating doses, which ranged from 800 mg/m2 to 2160 mg/m2, plus 500 mg/m2 DTIC. Plasma concentrations of gemcitabine and 2',2'-difluorodeoxyuridine, along with gemcitabine triphosphate (dFdCTP) levels in peripheral blood mononuclear cells (PBMCs), were evaluated during the course of treatment.

Protein kinase C theta is highly expressed in gastrointestinal stromal tumors but not in other mesenchymal neoplasias.

Purpose: Gastrointestinal stromal tumors (GIST) are a distinctive group of mesenchymal neoplasms of the gastrointestinal tract. The oncogene KIT has a central role in the pathogenesis of GIST, with c-kit receptor tyrosine kinase (KIT) protein expression being the gold standard in its diagnosis. The identification of GIST patients has become crucial, because the tyrosine kinase inhibitor Imatinib is effective in the treatment of this malignancy.

Cohesin component dynamics during meiotic prophase I in mammalian oocytes.

Cohesins are chromosomal proteins that form complexes involved in the maintenance of sister chromatid cohesion during division of somatic and germ cells. Three meiosis-specific cohesin subunits have been reported in mammals, REC8, STAG3 and SMC1 beta; their expression in mouse spermatocytes has also been described. Here we studied the localization of different meiotic and mitotic cohesin components during prophase I in human and murine female germ cells.

Secreted MMP9 promotes angiogenesis more efficiently than constitutive active MMP9 bound to the tumor cell surface.

Association of matrix metalloprotease 9 (MMP9) to the cell membrane is considered important in tumor growth and angiogenesis. To dissect this regulatory mechanism, we generated raft and non-raft MMP9 chimeras to force membrane expression in the MCF-7 human breast carcinoma cell line. MMP9 targeting to non-raft cell surface domains rendered a constitutive active membrane MMP9 form, suggesting a contribution by the lipid environment in MMP activation.

Sequential dose-dense doxorubicin and ifosfamide for advanced soft tissue sarcomas: a Phase II trial by the Spanish Group for Research on Sarcomas (GEIS).

Background: Combinations of high-dose ifosfamide (IF; 10-12 g/m2) plus doxorubicin (DX; 50-90 mg/m2) have been administered to patients with advanced soft tissue sarcoma (ASTS) in an attempt to improve therapeutic efficacy. Although these combination regimens appear to yield higher response rates than do standard-dose regimens, they also are associated with significant hematologic toxicity, despite the administration of hematopoietic growth factor support. As a potentially less toxic alternative, the authors designed a sequential, dose-dense schedule of DX and IF and explored its feasibility and toxicity, as well as patient compliance with the schedule, in a Phase II trial.

Activity of weekly irinotecan (CPT-11) in patients with advanced non-small cell lung cancer pretreated with platinum and taxanes.

Irinotecan (CPT-11), a semisynthetic derivative of camptothecin, is active in the treatment of non-small cell lung cancer (NSCLC). In this report we describe our experience with this drug when used as a single agent in patients with advanced NSCLC refractory to chemotherapy with platinum and taxanes. Nineteen NSCLC patients (thirteen males and six females; 53% adenocarcinoma and 26% squamous cell carcinoma) with a median age of 52 years (range 34-71) and a Karnofsky performance status of 60% (60-80%) were included in the study.

Irinotecan in the treatment of advanced colorectal cancer in patients pretreated with Fluorouracil-based chemotherapy: a study to determine recommendable therapeutic dosage.

Because no consensus exists regarding recommendable dose levels for irinotecan, an intrapatient dose escalation phase I-II study was initiated in previously treated patients with colorectal cancer. Survival was a secondary endpoint. Thirty-five consecutive patients with progressive disease after 5-fluorouracil-based chemotherapy were enrolled to receive irinotecan starting from 250 mg/m2/3 weeks and rising to currently used therapeutic doses.

Phase I/II study of gemcitabine and vinorelbine plus cisplatin in non-small cell lung cancer.

Purpose: We determine the maximum tolerated dose (MTD) and efficacy of gemcitabine plus vinorelbine combined with cisplatin in patients with non-small cell lung cancer (NSCLC).

Patients And Methods: Chemo naive patients with stage IIIA to IV non-small cell lung cancer received outpatient administration of gemcitabine 1,000 mg/m2 and vinorelbine 25 mg/m2 intravenously on days 1 and 8 every 21 days. Doses of gemcitabine and vinorelbine were escalated by 250 mg/m2 and 5 mg/m2, respectively, at each dose level.

STAG2 and Rad21 mammalian mitotic cohesins are implicated in meiosis.

STAG/SA proteins are specific cohesin complex subunits that maintain sister chromatid cohesion in mitosis and meiosis. Two members of this family, STAG1/SA1 and STAG2/SA2,double dagger are classified as mitotic cohesins, as they are found in human somatic cells and in Xenopus laevis as components of the cohesin(SA1) and cohesin(SA2) complexes, in which the shared subunits are Rad21/SCC1, SMC1 and SMC3 proteins. A recently reported third family member, STAG3, is germinal cell-specific and is a subunit of the meiotic cohesin complex.

Phase I/II study of gemcitabine plus vinorelbine in non-small cell lung cancer.

Background: Because gemcitabine and vinorelbine have demonstrated single-agent activity in non-small cell lung cancer (NSCLC), we conducted this phase I/II study to determine the maximum tolerated dose (MTD) and activity of these drugs combined.

Patients And Methods: Patients with inoperable or advanced NSCLC and no prior chemotherapy were treated with gemcitabine plus vinorelbine on days 1 and 8 every 21 days. The initial doses of gemcitabine 1,000 mg/m2 and vinorelbine 25 mg/m2 were escalated by 250 mg/m2 and 5 mg/m2, respectively, in separate patient cohorts until the MTD was established.