Neutrophil gelatinase-associated lipocalin (NGAL) predicts response to neoadjuvant chemotherapy and clinical outcome in primary human breast cancer

In our previous work we showed that NGAL, a protein involved in the regulation of proliferation and differentiation, is overexpressed in human breast cancer (BC) and predicts poor prognosis. In neoadjuvant chemotherapy (NACT) pathological complete response (pCR) is a predictor for outcome. The aim of this study was to evaluate NGAL as a predictor of response to NACT and to validate NGAL as a prognostic factor for clinical outcome in patients with primary BC. Immunohistochemistry was performed on tissue microarrays from 652 core biopsies from BC patients, who underwent NACT in the GeparTrio trial. NGAL expression and intensity was evaluated separately. NGAL was detected in 42.2% of the breast carcinomas in the cytoplasm. NGAL expression correlated with negative hormone receptor (HR) status, but not with other baseline parameters. NGAL expression did not correlate with pCR in the full population, however, NGAL expression and staining intensity were significantly associated with higher pCR rates in patients with positive HR status. In addition, strong NGAL expression correlated with higher pCR rates in node negative patients, patients with histological grade 1 or 2 tumors and a tumor size <40 mm. In univariate survival analysis, positive NGAL expression and strong staining intensity correlated with decreased disease-free survival (DFS) in the entire cohort and different subgroups, including HR positive patients. Similar correlations were found for intense staining and decreased overall survival (OS). In multivariate analysis, NGAL expression remained an independent prognostic factor for DFS. The results show that in low-risk subgroups, NGAL was found to be a predictive marker for pCR after NACT. Furthermore, NGAL could be validated as an independent prognostic factor for decreased DFS in primary human BC

Industrial biotechnology of Pseudomonas putida: advances and prospects

Pseudomonas putida is a Gram-negative, rod-shaped bacterium that can be encountered in diverse ecological habitats. This ubiquity is traced to its remarkably versatile metabolism, adapted to withstand physicochemical stress, and the capacity to thrive in harsh environments. Owing to these characteristics, there is a growing interest in this microbe for industrial use, and the corresponding research has made rapid progress in recent years. Hereby, strong drivers are the exploitation of cheap renewable feedstocks and waste streams to produce value-added chemicals and the steady progress in genetic strain engineering and systems biology understanding of this bacterium. Here, we summarize the recent advances and prospects in genetic engineering, systems and synthetic biology, and applications of P. putida as a cell factory

Prospective comparison of handheld ultrasound devices from different manufacturers with respect to B-scan quality and clinical significance for various abdominal sonography questions

Background: Handheld ultrasound (HHUS) devices have chiefly been deployed in emergency medicine, where they are considered a valid tool. The data situation is less clear in the case of internal questions in abdominal sonography. In our study, we investigate whether HHUS devices from different manufacturers differ in their B-scan quality, and whether any differences are relevant for the significance of an internal ultrasound examination. Method: The study incorporated eight HHUS devices from different manufacturers. Ultrasound videos of seven defined sonographic questions were recorded with all of the devices. The analogue recording of the same findings with a conventional high-end ultrasound (HEUS) device served as an evaluation criterion. Then, the corresponding findings were played side by side and evaluated by fourteen ultrasound experts using a point scale (5 points = very good; 1 point = insufficient). Results: The HHUS devices achieved relatively good results in terms of both the B-scan quality assessment and the ability to answer the clinical question, regardless of the manufacturer. One of the tested HHUS devices even achieved a significantly (p < 0.05) higher average points score in both the evaluation of B-scan quality and in the evaluation of clinical significance than the other devices. Regardless of the manufacturer, the HHUS devices performed best when determining the status/inferior vena cava volume and in the representation of ascites/free fluid. Conclusion: In various clinical abdominal sonography questions, HHUS systems can reliably reproduce findings, and are—while bearing their limitations in mind—an acceptable alternative to conventional HEUS systems. Irrespective of this, the present study demonstrated relevant differences in the B-scan quality of HHUS devices from different manufacturers

Biobased PET from lignin using an engineered cis, cis-muconate-producing Pseudomonas putida strain with superior robustness, energy and redox properties

Polyethylene terephthalate (PET), the most common synthetic polyester today, is largely produced from fossil resources, contributing to global warming. Consequently, sustainable sources must be developed to meet the increasing demand for this useful polymer. Here, we demonstrate a cascaded value chain that provides green PET from lignin, the world's most underutilized renewable, via fermentative production of cis, cis-muconate (MA) from lignin-based aromatics as a central step. Catechol, industrially the most relevant but apparently also a highly toxic lignin-related aromatic, strongly inhibited MA-producing Pseudomonas putida MA-1. Assessed by 13C metabolic flux analysis, the microbe substantially redirected its carbon core fluxes, resulting in enhanced NADPH supply for stress defense but causing additional ATP costs. The reconstruction of MA production in a genome-reduced P. putida chassis yielded novel producers with superior pathway fluxes and enhanced robustness to catechol and a wide range of other aromatics. Using the advanced producer P. putida MA-10 catechol, MA could be produced in a fed-batch process from catechol (plus glucose as additional growth substrate) up to an attractive titer of 74 g L-1 and a space-time-yield of 1.4 g L-1 h-1. In terms of co-consumed sugar, the further streamlined strain MA-11 achieved the highest yield of 1.4 mol MA (mol glucose)-1, providing a striking economic advantage. Following fermentative production, bio-based MA was purified and used to chemically synthetize the PET monomer terephthalic acid and the comonomer diethylene glycol terephthalic acid through five steps, which finally enabled the first green PET from lignin

Systems biology of electrogenic Pseudomonas putida - multi-omics insights and metabolic engineering for enhanced 2-ketogluconate production

Background Pseudomonas putida KT2440 has emerged as a promising host for industrial bioproduction. However, its strictly aerobic nature limits the scope of applications. Remarkably, this microbe exhibits high bioconversion efficiency when cultured in an anoxic bio-electrochemical system (BES), where the anode serves as the terminal electron acceptor instead of oxygen. This environment facilitates the synthesis of commercially attractive chemicals, including 2-ketogluconate (2KG). To better understand this interesting electrogenic phenotype, we studied the BES-cultured strain on a systems level through multi-omics analysis. Inspired by our findings, we constructed novel mutants aimed at improving 2KG production. Results When incubated on glucose, P. putida KT2440 did not grow but produced significant amounts of 2KG, along with minor amounts of gluconate, acetate, pyruvate, succinate, and lactate. 13C tracer studies demonstrated that these products are partially derived from biomass carbon, involving proteins and lipids. Over time, the cells exhibited global changes on both the transcriptomic and proteomic levels, including the shutdown of translation and cell motility, likely to conserve energy. These adaptations enabled the cells to maintain significant metabolic activity for several weeks. Acetate formation was shown to contribute to energy supply. Mutants deficient in acetate production demonstrated superior 2KG production in terms of titer, yield, and productivity. The ∆aldBI ∆aldBII double deletion mutant performed best, accumulating 2KG at twice the rate of the wild type and with an increased yield (0.96 mol/ mol). Conclusions By integrating transcriptomic, proteomic, and metabolomic analyses, this work provides the first systems biology insight into the electrogenic phenotype of P. putida KT2440. Adaptation to anoxic-electrogenic conditions involved coordinated changes in energy metabolism, enabling cells to sustain metabolic activity for extended periods. The metabolically engineered mutants are promising for enhanced 2KG production under these conditions. The attenuation of acetate synthesis represents the first systems biology-informed metabolic engineering strategy for enhanced 2KG production in P. putida. This non-growth anoxic-electrogenic mode expands our understanding of the interplay between growth, glucose phosphorylation, and glucose oxidation into gluconate and 2KG in P. putida

Development and Integration of DOPS as Formative Tests in Head and Neck Ultrasound Education : Proof of Concept Study for Exploration of Perceptions

In Germany, progress assessments in head and neck ultrasonography training have been carried out mainly theoretically and lack standardisation. Thus, quality assurance and comparisons between certified courses from various course providers are difficult. This study aimed to develop and integrate a direct observation of procedural skills (DOPS) in head and neck ultrasound education and explore the perceptions of both participants and examiners. Five DOPS tests oriented towards assessing basic skills were developed for certified head and neck ultrasound courses on national standards. DOPS tests were completed by 76 participants from basic and advanced ultrasound courses (n = 168 documented DOPS tests) and evaluated using a 7-point Likert scale. Ten examiners performed and evaluated the DOPS after detailed training. The variables of “general aspects” (6.0 Scale Points (SP) vs. 5.9 SP; p = 0.71), “test atmosphere” (6.3 SP vs. 6.4 SP; p = 0.92), and “test task setting” (6.2 SP vs. 5.9 SP; p = 0.12) were positively evaluated by all participants and examiners. There were no significant differences between a basic and advanced course in relation to the overall results of DOPS tests (p = 0.81). Regardless of the courses, there were significant differences in the total number of points achieved between individual DOPS tests. DOPS tests are accepted by participants and examiners as an assessment tool in head and neck ultrasound education. In view of the trend toward “competence-based” teaching, this type of test format should be applied and validated in the future

Combined PARP and Dual Topoisomerase Inhibition Potentiates Genome Instability and Cell Death in Ovarian Cancer

Although ovarian cancer is a rare disease, it constitutes the fifth leading cause of cancer death among women. It is of major importance to develop new therapeutic strategies to improve survival. Combining P8-D6, a novel dual topoisomerase inhibitor with exceptional anti-tumoral properties in ovarian cancer and compounds in preclinical research, and olaparib, a PARP inhibitor targeting DNA damage repair, is a promising approach. P8-D6 induces DNA damage that can be repaired by base excision repair or homologous recombination in which PARP plays a major role. This study analyzed benefits of combining P8-D6 and olaparib treatment in 2D and 3D cultures with ovarian cancer cells. Measurement of viability, cytotoxicity and caspase activity were used to assess therapy efficacy and to calculate the combination index (CI). Further DNA damage was quantified using the biomarkers RAD51 and γH2A.X. The combinational treatment led to an increased caspase activity and reduced viability. CI values partially show synergisms in combinations at 100 nM and 500 nM P8-D6. More DNA damage accumulated, and spheroids lost their membrane integrity due to the combinational treatment. While maintaining the same therapy efficacy as single-drug therapy, doses of P8-D6 and olaparib can be reduced in combinational treatments. Synergisms can be seen in some tested combinations. In summary, the combination therapy indicates benefits and acts synergistic at 100 nM and 500 nM P8-D6

Inhibition of ADAM17 increases the cytotoxic effect of cisplatin in cervical spheroids and organoids

Introduction: Cervical cancer represents one of the main causes of female, cancer-related mortality worldwide. The majority of cancers are caused by human papillomaviruses such as HPV16 and HPV18. As chemotherapeutic resistance to first-line platinum treatment is still a predominant clinical challenge in advanced cervical cancer, novel treatment options including combinatorial therapies are urgently required to overcome chemotherapeutic resistance. Inhibition of A Disintegrin And Metalloproteinase (ADAM)-family members, heavily involved in tumour progression of a vast range of solid tumours, strongly improved response to chemotherapeutic treatment in other tumour entities including ovarian cancer. Methods: We established two- and three-dimensional models derived from three traditional cervical cancer cell lines and ectocervical cancer-derived organoids. Following characterisation, these models were used to investigate their response to cisplatin treatment in the absence and presence of ADAM inhibitors using viability assays and automated live cell imaging. Results: The pivotal role of the metalloprotease ADAM17 driving chemotherapy resistance was detectable in all ectocervical cultures irrespective of the model system used, whereas ADAM10 inhibition was predominantly effective only in loosely aggregated spheroids. We showed prominent differences regarding treatment responses between 2D monolayers compared to 3D spheroid and 3D organoid model systems. Particularly, the organoid system, regarded as the closest representation of primary tumours, exhibited reliably the combinatorial effect of ADAM17 inhibition and cisplatin in all three individual donors. Discussion: As two- and three-dimensional models of the same cell lines differ in their responses to chemotherapy it is essential to validate treatment strategies in more advanced model systems representing the patient situation more realistically. Ectocervical organoids showed reliable results regarding treatment responses closely mimicking the primary tumours and could therefore serve as an important tool for personalized medicine in cervical cancer. These findings strengthen the role of ADAM17 as a potential novel target for combinatorial treatments to overcome chemoresistance in cervical cancer

Zur Interpretation von Energieszenarien

Szenarien haben sich in vielen Bereichen als Standardkonzept für das systematische „Nachdenken über die Zukunft“ etabliert. Besonders häufig sind sie dort anzutreffen, wo ein Bedarf nach Orientierung (weil Problemstellungen von übergreifender, gesellschaftlicher Bedeutung sind) sowie große Unsicherheiten über die Ausprägung wesentlicher Einflussgrößen bestehen und es unterschiedliches Wissen, verschiedene Meinungen und Ansichten zu integrieren gilt. All das trifft auf die zukünftige Energieversorgung zu. So sind Entscheidungen zur Ausgestaltung des Energiesystems mit weit in die Zukunft reichenden Festlegungen sowie erheblichen ökonomischen, ökologischen und sozialen Konsequenzen verbunden. Dies betrifft nicht nur Entscheidungen über die Erforschung und Entwicklung neuer Technologien, sondern gerade auch Entscheidungen über den Ausbau konkreter Technologien wie etwa die Errichtung bestimmte Kraftwerke. Schließlich geht es dabei nicht nur um Investitionen, die sich erst nach Jahrzehnten amortisieren, sondern unter Umständen auch um langfristige Konsequenzen, die weit über die Betriebsdauer der Anlage hinausreichen – man denke etwa an die Lagerung des radioaktiven Abfalls aus Kernkraftwerken

ADAM17 Inhibition Increases the Impact of Cisplatin Treatment in Ovarian Cancer Spheroids

Chemotherapy resistance is a major challenge in ovarian cancer (OvCa). Thus, novel treatment combinations are highly warranted. However, many promising drug candidates tested in two-dimensional (2D) cell culture have not proved successful in the clinic. For this reason, we analyzed our drug combination not only in monolayers but also in three-dimensional (3D) tumor spheroids. One potential therapeutic target for OvCa is A disintegrin and metalloprotease 17 (ADAM17). ADAM17 can be activated by chemotherapeutics, which leads to enhanced tumor growth due to concomitant substrate cleavage. Therefore, blocking ADAM17 during chemotherapy may overcome resistance. Here, we tested the effect of the ADAM17 inhibitor GW280264X in combination with cisplatin on ovarian cancer cells in 2D and 3D. In 2D, the effect on five cell lines was analyzed with two readouts. Three of these cell lines formed dense aggregates or spheroids (HEY, SKOV-3, and OVCAR-8) in 3D and the treatment effect was analyzed with a multicontent readout (cytotoxicity, viability, and caspase3/7 activation). We tested the combined therapy on tumor spheroids derived from primary patient cells. In 2D, we found a significant reduction in the half minimal (50%) inhibitory concentration (IC50) value of the combined treatment (GW280264X plus cisplatin) in comparison with cisplatin monotherapy in all five cell lines with both 2D readout assays (viability and caspase activation). In contrast, the combined treatment only showed an IC50 reduction in HEY and OVCAR-8 3D tumor spheroid models using caspase3/7 activity or CelltoxTM Green as the readout. Finally, we found an improved effect of GW280264X with cisplatin in tumor spheroids derived from patient samples. In summary, we demonstrate that ADAM17 inhibition is a promising treatment strategy in ovarian cancer