Improved therapeutic entities derived from known generics as an unexplored source of innovative drug products

With a New Drug Application (NDA) innovative drug therapies are reaching the market in a specific dosage form for one or more clinically proven indications of which after expiration of the patent or the data exclusivity copies are launched using Abbreviated New Drug Applications (ANDA). Advanced therapies that emerged from launched molecules during their product life-cycle have gained considerable attention as clinical practice provides evidence for additional therapeutic values, patient centric delivery systems show improved therapeutic outcomes or emerging technologies offer efficiency gains in manufacturing or access to emerging markets. The USA and European regulatory framework has set reasonable regulations in place for these "Supergenerics" or "hybrid" applications. While these regulations are relatively recent the pharmaceutical industry is just starting to use this route for their product development and life-cycle management. From a clinical perspective the potential for advanced product development have been demonstrated. Yet, there is still a lag of common understanding between the different stakeholders regarding the development, application process and commercial incentive in developing enhanced therapeutic entities based on existing drug products for the market.     

2D and 3D collagen and fibrin biopolymers promote specific ECM and integrin gene expression by vascular smooth muscle cells

Collagen Type I and fibrin are polymeric proteins commonly used in the field of regenerative medicine as the foundational matrix of engineered tissues. We examined the response of vascular smooth muscle cells (VSMC) to both two-dimensional (2D) substrates as well as three-dimensional (3D) matrices of these biopolymers. Pure collagen Type I, pure fibrin and composite matrices consisting of 1:1 mixtures of collagen and fibrin were studied. Relative gene expression of three ECM molecules (collagen Type I and III, and tropoelastin) and three integrin subunits (integrins alpha1, beta1 and beta3) was determined over 7 days in culture using quantitative RT-PCR. Expression of all of these marker genes was up-regulated in 3D matrices, relative to 2D substrates. Tropoelastin, integrin alpha1 and integrin beta1 were highest in collagen matrices, while collagen III and integrin beta3 expression were highest in pure fibrin, and collagen I expression was highest in the collagen-fibrin composite materials. Both the compositional and temporal expression patterns of these specific ECM-related genes were suggestive of a wound healing response. These results illuminate the short-term responses of VSMC to 2D and 3D biopolymer matrices, and have relevance to tissue engineering and cardiovascular biology.     

Dopamine as additive to cold preservation solution improves postischemic integrity of the liver

Dopamine pretreatment has been used to confer protection against cellular injury following hypothermia or anoxia, especially in vascular endothelial cells. Ischemia/reperfusion‐associated tissue alterations still represent a major drawback in liver transplantation. The present study was aimed to investigate the effect of dopamine as an ex vivo adjunct, added to the cold storage solution, on cold preservation of the liver. Rat livers were excised 30 min after cardiac arrest, flushed with preservation solution and cold stored for 18 h. Dopamine (10, 50 or 100 μm ) was added to the preservation solution in other livers. Organ viability was evaluated by 120 min of warm reperfusion in vitro (n = 6, resp.). Dopamine induced a dose related up to fourfold (at 50 μm ) reduction in parenchymal (ALT, LDH) and mitochondrial (GLDH) enzyme release and significantly reduced histologic signs of tissue injury. Bile production and tissue ATP was doubled by dopamine. On the molecular level, dopamine enhanced postischemic phosphorylation of protein kinase A and p42/44 MAP kinase. Inhibition of cAMP‐PKA pathway by simultaneous application of RP‐cAMPs had no effect on P42/44 phosphorylation, or functional recovery of dopamine‐treated grafts. Dopamine supplementation of the flush‐out solution appears as a simple way for ex vivo augmentation of liver viability during preservation, not mediated via the catecholamine‐cAMP signal cascade.     

Allgemeine Pathologie und pathologische Anatomie

International Journal of Legal Medicine -     

Zur Physiologie der Blutgefässchirurgie

Ohne Zusammenfassung     

Targeting of NADPH oxidase in vitro and in vivo suppresses fibroblast activation and experimental skin fibrosis

Although there is increasing evidence that oxidative stress is involved in collagen synthesis and myofibroblast activation, the NADPH oxidase (Nox) system is incompletely investigated in the context of human dermal fibroblasts (HDFs) and skin fibrosis. Using the pan‐Nox inhibitor diphenyleneiodonium (DPI) as an initial tool, we show that gene expression of collagen type I, α‐smooth muscle actin (α‐SMA) and fibronectin 1 is suppressed in HDFs. Detailed expression analysis of all Nox isoforms and adaptors revealed expression of RNA and protein expression of Nox4, p22^phox and Poldip2 but neither Nox1 nor Nox2. Nox4 could be immunolocalized to the endoplasmic reticulum. Importantly, TGF‐β₁ had a dose‐ and time‐dependent upregulating effect on NADH activity and Nox4 gene expression in HDFs. Genetic silencing of Nox4 as demonstrated by siRNA in HDFs as well as in murine fibroblasts established from Nox4 knockout mice confirmed that TGF‐β₁‐mediated collagen type I gene, α‐SMA and fibronectin 1 gene expressions were Nox4‐dependent. This TGF‐β₁ effect was mediated by Smad3 as shown by in silico promoter analysis, pharmacological inhibition and gene silencing of Smad3. The relevance of these findings is highlighted in the bleomycin‐induced scleroderma mouse model. DPI treatment attenuated skin fibrosis and myofibroblast activation. Moreover, Nox4 knockdown by siRNA reduced skin collagen synthesis, α‐SMA and fibronectin 1 expression in vivo. Finally, analyses of HDFs from patients with systemic sclerosis confirmed the expression of Nox4 and its adaptors, whereas Nox1 and Nox2 were not detectable. Our findings indicate that Nox4 targeting is a promising future treatment for fibrotic skin diseases.     

Die Beziehung zwischen der funktionellen Totraumventilation, der Atemform und der CO2-Abgabe beim ruhenden und arbeitenden Menschen

Zusammenfassung In Untersuchungen an vier gesunden männlichen Versuchspersonen wurden der arterielle CO₂-Druck (nachAstrup), die Ventilation, die Atemfrequenz, die CO₂-Abgabe und O₂-Aufnahme bei Ruhe und Leistungsstufen bis 15 mkp/sec am Fahrradergometer gemessen. Berechnet wurden der funktioneile Totraum und das Verhältnis von alveolärer Ventilation zur Gesamtventilation (alveolärer Wirkungsgrad). Es zeigte sich, daß der funktionelle Totraum unabhängig von der Leistung bei allen Versuchspersonen jeweils um den gleichen Mittelwert streute. Der alveoläre Wirkungsgrad nahm mit steigender Atemfrequenz ab, stieg jedoch mit zunehmender CO₂-Abgabe an. Bei Arbeit wird die Vergrößerung der Totraumventilation, die durch Erhöhung der Atemfrequenz bewirkt wird, durch die höhere CO₂-Abgabe kompensiert.

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Summary The arterial CO₂ pressure, determined byAstrup's method, the ventilation, the respiratory rate, the oxygen consumption and the CO₂ output were measured in four healthy young men during rest and exercise on a bicycle ergometer up to 15 mkp/sec. The functional dead space and the ratio between alveolar and total ventilation (alveolar efficiency) were calculated as functions of performance. The functional dead space of each subject was independend of the performance.A considerable scatter was found in single measurements. There was an increase in alveolar efficiency, when the CO₂ output rose, but a decrease with increasing respiratory frequency. During exercise the augmentation of dead space ventilation caused by the increase respiratory rate is compensated by the increased CO₂ output.

     

Prediction of individual oxygen uptake on-step transients from frequency responses

Power-oxygen uptake ( ) frequency responses can be used to predict responses to arbitrary exercise intensity patterns. It is still an open question for which range of exercise intensities such computed response patterns yield valid predictions. In the present study, we determined the power- frequency response of nine sports students by means of pseudo-randomised switching between 20 W and 80 W during upright and supine cycle exercise. Starting from a baseline of 20 W each subject also performed sustained step increases to 40 W, 80 W, 120 W, and 160 W in both positions. The individual step responses were then compared with the expected time-courses predicted on the basis of the individual frequency responses. The comparison showed a close agreement for the 20 W–40 W and 20 W–80 W steps in both positions. With larger step amplitudes the kinetics became increasingly slower than the predicted time course in both positions. During additional ramp tests (10 W · 30 s^–1) whole blood lactic acid concentration [1a^–]_b tended to be higher in the supine position at exercise intensities higher than 160 W. The mean power at 4 mmol · 1^–1 [la^–]_b amounted to 234 (SD 32) W and 253 (SD 44) W (P<5%) in the supine and the upright position, respectively. The maximal oxygen uptake relative to body mass was not found to be significantly different [upright, mean 57 (SD 10) ml · (min · kg)^–1;supine, mean 54 (SD 10) ml · (min · kg)]. These findings would suggest that for a range of mild exercise intensities kinetics are not appreciably influenced by the step amplitude or by cardiovascular changes associated with the upright and the supine position.