Regulatory and health technology assessment advice on postlicensing and postlaunch evidence generation is a foundation for lifecycle data collection for medicines.

The understanding of the benefit risk profile, and relative effectiveness of a new medicinal product, are initially established in a circumscribed patient population through clinical trials. There may be uncertainties associated with the new medicinal product that cannot be, or do not need to be resolved before launch. Postlicensing or postlaunch evidence generation (PLEG) is a term for evidence generated after the licensure or launch of a medicinal product to address these remaining uncertainties. PLEG is thus part of the continuum of evidence development for a medicinal product, complementing earlier evidence, facilitating further elucidation of a product's benefit/risk profile, value proposition, and/or exploring broader aspects of disease management and provision of healthcare. PLEG plays a role in regulatory decision making, not only in the European Union but also in other jurisdictions including the USA and Japan. PLEG is also relevant for downstream decision-making by health technology assessment bodies and payers. PLEG comprises studies of different designs, based on data collected in observational or experimental settings. Experience to date in the European Union has indicated a need for improvements in PLEG. Improvements in design and research efficiency of PLEG could be addressed through more systematic pursuance of Scientific Advice on PLEG with single or multiple decision makers. To date, limited information has been available on the rationale, process or timing for seeking PLEG advice from regulators or health technology assessment bodies. This article sets out to address these issues and to encourage further uptake of PLEG advice.

Regulatory Pathways Supporting Expedited Drug Development and Approval in ICH Member Countries.

Regulators and pharmaceutical companies across the world are intensifying efforts to get increasingly complex and innovative drugs to patients with high unmet medical need in the shortest possible time frame. This article reviews pathways to expedite drug development and approval available in member countries of the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use and Australia. It is concluded that the increasing availability of expedited regulatory pathways and associated modernisation of regulatory systems changes the current regulatory paradigm and requires sponsors to rethink drug development and regulatory strategy. A transformation of the current sequence of regulatory submissions, favouring those countries/collaborations that are best regulatory equipped to make innovative medical need drugs available to patients in the shortest time frame is imminent.

The role of the cardiac Na+/Ca2+ exchanger in reverse remodeling: relevance for LVAD-recovery.

Different strategies can, at least in certain conditions, prevent or reverse myocardial remodeling due to heart failure and induce myocardial functional improvement. Na+/Ca2+ exchanger (NCX) is considered a major player in the pathophysiology of heart failure but its role in reverse remodeling is unknown. A combination of mechanical unloading by left ventricular assist devices (LVADs) and pharmacological therapy has been shown to induce clinical recovery in a limited number of patients with end-stage heart failure. In myocytes isolated from these patients we found that, after LVAD treatment, NCX1/SERCA2a mRNA was 38% higher than at device implant. We studied the ability of NCX to extrude Ca2+ during caffeine-induced SR Ca2+ release in isolated ventricular myocytes from these patients. The time constant of decline was slower in heart failure. In myocytes from patients with clinical recovery following mechanical and pharmacological treatment, NCX1-mediated Ca2+ extrusion was faster compared with myocytes from patient who, despite identical treatment, did not recover. We propose that increased NCX function may be associated with reverse remodeling in patients and that factors that regulate NCX function (i.e., phosphorylation or intracellular [Na+]) other than NCX expression levels alone, may have detrimental consequences on cardiac function.

Sp1 and Sp3 transcription factors are required for trans-activation of the human SERCA2 promoter in cardiomyocytes.

OBJECTIVES: The sarcoplasmic reticulum (SR) Ca(2+) ATPase (SERCA) is essential to the removal of cytosolic calcium following cardiac contraction, and its abundance and activity are significantly altered during perinatal development and in failing myocardium. The objective of the current study was to identify cis regulatory elements and nuclear transcription factors responsible for transactivating SERCA2 gene expression in cardiomyocytes. METHODS: Primary cultures of neonatal rat ventricular myocytes were transiently transfected with luciferase (LUX) reporter gene constructs containing deletions of the SERCA2 promoter or which harbored mutations in consensus Sp1 transcription factor binding sites. Cotransfection assays, electrophoretic mobility shift, and supershift assays were also performed to delineate the regulatory role of specific transcription factors. RESULTS: We identified a putative AP-1-like element and a consensus Egr-1 binding site, but neither Egr-1 nor 12-O-tetradecanoylphorbol 13-acetate (TPA) significantly modified human SERCA2 promoter activity in vitro. Maximal activity of the SERCA2 promoter required the proximal 177 bp, and strong activation was observed with a 125-bp construct, within which an Sp1 site and a CAAT box were important. Mutation analysis also revealed the importance of two Sp1 sites between -125 and -200. Sp1 and Sp3 transcription factors were subsequently identified to bind to oligonucleotide probes corresponding to only the two most proximal Sp1 sites. CONCLUSIONS: These studies provide direct evidence that regulation of human SERCA2 gene expression in cardiomyocytes depends on transactivation events elicited by Sp1 and Sp3 transcription factors.

Increased expression of extracellular matrix regulators TIMP1 and MMP1 in deteriorating heart failure.

BACKGROUND: The authors previously identified and compared alterations in gene expression in the myocardia of patients with deteriorating heart failure who underwent left ventricular assist device (LVAD) implantation with those of patients with stable end-stage failure (ESF). We hypothesized that matrix metalloproteinases (MMPs) and their endogenous inhibitors, the tissue inhibitors of MMPs (TIMPs), would be implicated in the mechanisms that underlie deteriorating heart failure. METHODS: Gridded macro-array filters were used to provide a broad overview of MMP and TIMP mRNA expression in heart failure. Precise mRNA levels of TIMP1, MMP1, and beta-spectrin were determined using quantitative real-time reverse-transcription polymerase chain reaction (RT-PCR) of myocardial samples from 27 patients with deteriorating heart failure who underwent LVAD implantation, from 17 patients with stable ESF who underwent elective heart transplantation, and from 28 donor organs with good hemodynamic function. RESULTS: Gridded macro-arrays analysis of pooled failing heart samples determined that TIMP1 mRNA was the most readily detectable TIMP in failing myocardium. Quantitative RT-PCR showed that expression levels in individual patients were similar in patients with stable ESF (1.00 +/- 0.24, n = 17) and in donor organ samples (1.49 +/- 0.22, n = 28) but were significantly increased in the deteriorating heart failure group (5.38 +/- 0.32, n = 26, p < 0.0001 compared with patients with ESF). Similarly, MMP1 levels did not differ between donor and ESF groups but increased in the deteriorating failure group (6.04 +/- 0.50, n = 27, p < 0.001 compared with the ESF group). Levels of beta-II spectrin were the same in all 3 groups. Both TIMP1 and MMP1 showed positive correlation with each other and with previously determined levels of mRNA for both interleukin-1beta (IL-1beta) and IL-6 in this patient series when considering all patients individually, but neither correlated with tumor necrosis factor alpha. CONCLUSIONS: Patients with deteriorating heart failure have increased expression of TIMP1 and MMP1 mRNA. Correlation with pro-inflammatory cytokines suggests common pathways of regulation and potential activation by IL-6 and IL1-beta.

The slow skeletal muscle troponin T gene is expressed in developing and diseased human heart.

Cardiac muscle development is characterised by the activation of contractile protein genes and subsequent modulation of expression resulting, ultimately, in the formation of a mature four-chambered organ. Myocardial gene expression is also altered in the adult in response to pathological stimuli and this is thought to contribute to the altered contractile characteristics of the diseased heart. We have examined the expression of the slow skeletal troponin T (TnT) gene in the human heart during development and in disease using whole mount in situ hybridisation and real-time quantitative (TaqMan) polymerase chain reaction (PCR). Slow skeletal TnT mRNA shows transitory and regional expression in the early foetal heart, which occurs at different times in atria and ventricles. In ventricular myocardium, expression is seen in the outer epicardial layer at a time when the coronary circulation is being established. Expression was detected at low levels in the adult human heart and was significantly increased in end-stage heart failure. Similarly, expression was readily detectable during early rat heart development and was up-regulated in pressure overload hypertrophy in adult. Together these data show for the first time that slow skeletal TnT mRNA is readily detectable during early human heart development. They further suggest that slow skeletal TnT may be responsive to myocardial stress and that elevated levels may contribute to myocardial dysfunction in adult disease.

The slow skeletal muscle troponin T gene is expressed in developing and diseased human heart.

Cardiac muscle development is characterised by the activation of contractile protein genes and subsequent modulation of expression resulting, ultimately, in the formation of a mature four-chambered organ. Myocardial gene expression is also altered in the adult in response to pathological stimuli and this is thought to contribute to the altered contractile characteristics of the diseased heart. We have examined the expression of the slow skeletal troponin T (TnT) gene in the human heart during development and in disease using whole mount in situ hybridisation and real-time quantitative (TaqMan) polymerase chain reaction (PCR). Slow skeletal TnT mRNA shows transitory and regional expression in the early foetal heart, which occurs at different times in atria and ventricles. In ventricular myocardium, expression is seen in the outer epicardial layer at a time when the coronary circulation is being established. Expression was detected at low levels in the adult human heart and was significantly increased in end-stage heart failure. Similarly, expression was readily detectable during early rat heart development and was up-regulated in pressure overload hypertrophy in adult. Together these data show for the first time that slow skeletal TnT mRNA is readily detectable during early human heart development. They further suggest that slow skeletal TnT may be responsive to myocardial stress and that elevated levels may contribute to myocardial dysfunction in adult disease. (Mol Cell Biochem 263: 91-97, 2004).

Changes in sarcolemmal Ca entry and sarcoplasmic reticulum Ca content in ventricular myocytes from patients with end-stage heart failure following myocardial recovery after combined pharmacological and ventricular assist device therapy.

AIMS: Support with left ventricular assist devices (LVAD) improves cardiac performance in patients with end-stage heart failure. In some cases this strategy, combined with pharmacological treatment, has led to a clinical improvement which remained after LVAD explant. This study defines changes in Ca handling at the cellular level in failing left ventricular tissue taken at LVAD implant (LVAD core) and LVAD removal (post-LVAD). METHODS AND RESULTS: We studied cell size and Ca regulation in enzymatically dissociated cardiac myocytes. We used confocal microscopy and electrophysiological techniques to investigate the SR Ca content and major Ca movements across the sarcolemma during the action potential. We firstly recorded a significant reduction in cell capacitance and cell volume consistent with regression of cellular hypertrophy in post-LVAD myocytes compared with LVAD core myocytes. Ca entry via sarcolemmal Ca channels during the action potential using action potential voltage-clamping was significantly increased in post-LVAD myocytes compared with LVAD cores myocytes. Finally, SR Ca content (assessed by integrating the caffeine-induced Na/Ca exchanger transient inward current) in post-LVAD myocytes was also significantly increased compared with LVAD cores myocytes. CONCLUSIONS: These results show that in myocytes from patients after LVAD support there is more Ca entry to trigger Ca release and more SR Ca content, leading to improved contractile function.