Scientific advances in diabetes: the impact of the innovative medicines initiative

Tese de mestrado, Regulação e Avaliação do Medicamento e Produtos de Saúde, 2020, Universidade de Lisboa, Faculdade de Farmácia.A Iniciativa sobre Medicamentos Inovadores é uma parceria público-privada entre a Comunidade Europeia, representada pela Comissão Europeia, e a Indústria Farmacêutica, representada pela Federação Europeia da Indústria Farmacêutica. Esta Iniciativa de Tecnologia Conjunta tem como objetivos acelerar o processo de investigação e desenvolvimento de medicamentos inovadores, bem como gerar novos conhecimentos científicos que promovam a integração da medicina personalizada nas doenças prioritárias definidas pela Organização Mundial de Saúde. Atualmente, no âmbito desta iniciativa foram estabelecidos dois programas, sendo que o primeiro (IMI1) decorreu entre 2008 e 2013 e teve um orçamento de 2 mil milhões de euros, enquanto que o segundo programa (IMI2) está em decurso desde 2014 e terminará em 2020 e o orçamento disponibilizado foi de 3.276 mil milhões de euros. A diabetes mellitus é uma das doenças prioritárias indicadas pela Organização Mundial de Saúde alvo de financiamento pelos programas IMI. As principais justificações para este facto prendem-se com os dados epidemiológicos da doença. No decorrer dos anos, verificou-se um aumento exponencial da taxa de prevalência desta doença a nível mundial. Esta evidência é suportada pelo facto de, entre o período de 1980 e 2014, esta taxa ter sofrido um aumento de 4.7% para 8.7%, o quadruplo do valor, em adultos com idade igual ou superior a 18 anos e também por as estimativas a 20 anos, realizadas pela Organização Mundial de Saúde, indicarem que o número total de casos existentes corresponderá a mais de 20% da população universal. Simultaneamente, constatou-se um crescimento progressivo tanto da taxa de mortalidade por diabetes bem como dos custos de saúde acarretados por esta doença. No que diz respeito à taxa de mortalidade, em 2016, a diabetes foi considerada a sétima principal causa de morte no mundo. Em termos de impacto económico, a diabetes e as complicações decorrentes desta doença, como é o caso das doenças cardiovasculares, nefropatia diabética e retinopatia diabética, impõem um grande peso económico para os sistemas de saúde. A nível mundial, os custos anuais provocados pela diabetes, entre o ano de 2007 e 2019, aumentaram de 232 mil milhões de dólares para 760 mil milhões de dólares, o que equivale a incremento de 528 mil milhões de dólares em 12 anos. Na área da Diabetes, o principal objetivo dos programas IMI1 e IMI2 é o de reduzir a tendência crescente observada na taxa de prevalência desta doença. De forma a atingir esta meta, os dois programas supramencionados primaram o financiamento de projetos cujo intuito consistia no desenvolvimento do conhecimento, medicamentos, métodos, ferramentas e modelos que facilitassem a implementação da medicina personalizada, como modelo de prática médica corrente, em doentes com diabetes. Até outubro de 2019, os programas IMI financiaram treze projetos para a área da Diabetes & Doenças Metabólicas, nomeadamente SUMMIT, IMIDIA, DIRECT, StemBANCC, EMIF, EBiSC, INNODIA, RHAPSODY, BEAT-DKD, LITMUS, Hypo-RESOLVE, IM2PACT e CARDIATEAM. Entre estes, o INNODIA tinha como objetivo a diabetes tipo 1, o DIRECT, EMIF e RHAPSODY tinham como foco a diabetes tipo 2, o SUMMIT, BEAT-DKD, LITMUS, Hypo-RESOLVE e CARDIATEAM estavam associados às complicações da diabetes e os restantes projetos, o StemBANCC, EBiSC, IMIDIA e IMI2PACT, estavam orientados para o desenvolvimento da vertente científica. Em geral, um investimento monetário total na ordem dos €447 249 438 foi realizado pelo IMI na área da Diabetes. Todavia, a deteção da lacuna existente na integração dos resultados produzidos pelos diferentes projetos, impulsionou a elaboração da presente dissertação intitulada de “Scientific Advances in Diabetes: The Impact of the Innovative Medicines Initiative”, ou seja, Avanços Científicos na Área da Diabetes: Impacto da Iniciativa sobre Medicamentos Inovadores. Os principais objetivos estabelecidos para esta dissertação foram os de recolher os artigos publicados pelos projetos financiados e sistematizá-los nos eixos de investigação definidos na agenda estratégica do programa IMI2, mais concretamente: 1) identificação de alvos e biomarcadores, 2) novos paradigmas de ensaios clínicos, 3) medicamentos inovadores e 4) programas de adesão terapêutica centrados nos doentes. A metodologia de investigação aplicada nesta dissertação consistiu numa revisão de literatura, tendo-se utilizado como fontes de dados as páginas eletrónicas oficiais de cada projeto, o contacto com os coordenadores e co-coordenadores dos projetos e a base de dados europeia Cordis. No geral, um total de 662 citações foram identificadas, das quais 185 foram incluídas na análise realizada neste trabalho. Através da sistematização e integração dos artigos recolhidos nos projetos financiados pelo IMI, averiguou-se que para o eixo de identificação de alvos e biomarcadores, os outcomes relevantes responderam a cinco das recomendações definidas na agenda estratégica do programa IMI2, nomeadamente: 1) identificar e validar marcadores biológicos, ferramentas e ensaios, 2) identificar as determinantes que justificam a variabilidade interindividual, 3) compreender os mecanismos moleculares subjacentes à doença, 4) desenvolver uma plataforma de ensaios pré-clínicos e 5) estabelecer modelos de sistemas. De um modo geral, um vasto número de biomarcadores, ferramentas, fatores responsáveis pela heterogeneidade da população, incluindo marcadores genéticos, e mecanismos relevantes foram identificados para a diabetes tipo 1 pelo INNODIA, para a diabetes tipo 2 pelo SUMMIT, IMIDIA, DIRECT e EMIF, para as células beta pancreáticas pelo IMIDIA e RHAPSODY, para a nefropatia diabética pelo SUMMIT e BEAT-DKD, e para as doenças cardiovasculares e retinopatia diabética pelo SUMMIT. Suplementarmente, um conjunto de ferramentas e ensaios foram desenvolvidos pelos projetos StemBANCC, EBiSC e IMIDIA com o intuito de impulsionar avanços na área investigacional. Ainda neste eixo foram propostos dois modelos de estratificação dos doentes, um relativo ao controlo glicémico em doentes com diabetes tipo 1 estabelecido pelo INNODIA e outro correspondente à identificação dos subtipos de doentes com diabetes desenvolvido pela parceria BEAT-DKD/RHAPSODY. Relativamente ao eixo de ensaios clínicos, os dados analisados compreendiam propostas de novos parâmetros clínicos e de desenhos de ensaios, sendo que estes resultados visavam espelharem com maior precisão as características da subpopulação com diabetes em teste. Os dados incluídos neste eixo foram obtidos a partir dos projetos SUMMIT, DIRECT e BEAT-DKD. No que concerne ao eixo de medicamentos inovadores, as informações recolhidas dos artigos publicados pelos projetos SUMMIT, IMIDIA, DIRECT, StemBANNC, EMIF, INNODIA e BEAT-DKD consistiam na identificação de novos potenciais alvos terapêuticos bem como no desenvolvimento de novos agentes terapêuticos, ambos com a finalidade de tratar ou prevenir tanto a diabetes como as complicações associadas a esta doença. Foi ainda proposta uma nova abordagem de produção de células estaminais pluripotentes humanas em larga escala pelo StemBANCC. No que tange aos programas de maximização de resultados de saúde benéficos centrados no doente com diabetes, dois novos modelos preditivos foram desenvolvidos e validados pelo projeto DIRECT, permitindo a sua utilização como ferramentas de diagnóstico por médicos especialistas. Adicionalmente, esta dissertação tem como objetivo apresentar uma proposta de visão de complementaridade entre os treze projetos financiados pelo IMI, realçando as possíveis estratégias a adotar para a integração da medicina personalizada na prática clínica. Esta abordagem engloba a criação de indicadores biológicos e genéticos que facilitem a identificação dos indivíduos com risco elevado de desenvolver diabetes, a inclusão de ferramentas que possibilitem o diagnóstico precoce dos doentes e, por último, a seleção do tratamento apropriado às características do indivíduo, ou seja o que evidencie ser mais eficaz e seguro, suportado em modelos de estratificação de doentes, tentando desta forma retardar a progressão da doença, assim como prevenir o desenvolvimento das complicações relacionadas com a progressão da doença.Innovative Medicines Initiative (IMI) is a public-private partnership between the European Community, represented by the European Commission, and the European Federation of Pharmaceutical Industries and Associations. This joint undertaking aims at accelerating the medicines development process and generating new scientific knowledge to promote the implementation of personalized medicine for priority diseases established by the World Health Organization. Currently, two IMI programmes have been undertaken, the first one (IMI1) was carried out from 2008 until 2013 and had a budget of €2 billion, and the second one (IMI2) was developed from 2014 up to 2020 and the budget committed was up to €3.276 billion. Diabetes Mellitus is one of the World Health Organization’s priority diseases under research by the IMI programmes, mainly due to the exponential increase of its global prevalence over the years. Between 1980 and 2014, this rate quadrupled from 4.7% to 8.7% in adults aged 18 years and older and the 20 years- World Health Organization’s projections indicate that it could reach more than 20% of the population. Simultaneously, the mortality rate and the healthcare costs associated with diabetes have been increasing. Regarding mortality, diabetes was the seventh leading cause of death in 2016. In terms of economic impact, currently, diabetes and its related complications, such as cardiovascular diseases, diabetic kidney disease and diabetic retinopathy, represent a significant economic burden on the healthcare systems. Worldwide, the estimated annual costs of diabetes have increased from $232 billion to$760 billion, between 2007 and 2019. In the Diabetes field, the main aim of IMI1 and IMI2 programmes is to shorten the prevalence of this disease, through the development of knowledge and methods that enable the implementation of personalized treatment for diabetic patients. Up to October of 2019, thirteen projects were funded by IMI for Diabetes & Metabolic disorders, more precisely SUMMIT, IMIDIA, DIRECT, StemBANCC, EMIF, EBiSC, INNODIA, RHAPSODY, BEAT-DKD, LITMUS, Hypo-RESOLVE, IM2PACT, and CARDIATEAM. Of these, INNODIA aimed at type 1 diabetes, DIRECT, EMIF and RHAPSODY were focused on type 2 diabetes, SUMMIT, BEAT-DKD, LITMUS, Hypo- RESOLVE and CARDIATEAM were related to complications of diabetes, and the remaining projects, namely StemBANCC, EBiSC, IMIDIA and IMI2PACT, were directed to scientific research. In general, a total of €447 249 438 was spent by IMI in the area of Diabetes. However, there is a substantial lack of integration of achievements between the different projects, which prompted the development of this dissertation: “Scientific Advances in Diabetes: The Impact of the Innovative Medicines Initiative”. This dissertation’ objectives were to collect the data of the funded-projects and integrate them into the following research axes: 1) target and biomarker identification, 2) innovative clinical trials paradigms, 3) innovative medicines, and 4) patient-tailored adherence programmes. The research methodology applied was a literature review and the data sources used were the official project’s websites, contacts with the project’s coordinators and co-coordinator and the CORDIS database. From the 662 citations identified, 185 were included. Through the integration of the data collected from IMI-funded projects, it was verified that for Target and Biomarker identification, the main achievements were in order to 1) identify and validate biological markers, tools and assays, 2) identify determinants of inter-individual variability, 3) understand the molecular mechanisms underlying the disease, 4) develop a platform of pre-clinical assays, and 5) develop systems’ models. Therefore, several biomarkers, tools, inter-individual variability factors, including genetic markers, and relevant pathways were proposed for type 1 diabetes by INNODIA, for type 2 diabetes by SUMMIT, IMIDIA, DIRECT and EMIF, for pancreatic β-cells by IMIDIA and RHAPSODY, for diabetic kidney disease by SUMMIT and BEAT-DKD, and for cardiovascular diseases and diabetic retinopathy by SUMMIT. Moreover, new tools and assays to improve research field were developed by StemBANCC, EBiSC and IMIDIA. Also, two models for patients’ stratification were proposed, one related to glycaemic control in patients with type 1 diabetes established by INNODIA, and another corresponding to the identification of subtypes of diabetes patients developed by BEAT-DKD/RHAPSODY. Regarding the clinical trials, the data collected SUMMIT, DIRECT and BEAT-DKD corresponds to new clinical endpoints and trial designs to accurately reflect the characteristics of the diabetic subpopulation under test. In terms of innovative medicines, information retrieved by SUMMIT, IMIDIA, DIRECT, StemBANNC, EMIF, INNODIA and BEAT-DKD consists on the identification of new therapeutic targets and the development of agents with the purpose of treatment and prevent diabetes and its related complications. Furthermore, a new approach for the large-scale production of human pluripotent stem cells was proposed by StemBANCC. Concerning the maximization of beneficial health patient-centred outcomes, two novel predictive models were developed and validated by DIRECT for diabetes to be used as screening tools by doctors. In addition, this dissertation intends to present a joint vision of the IMI-projects with strategies for integrating personalized medicine into healthcare practice. This approach involves the creation of biological and genetic indicators that can be used to identify individuals at high risk of developing diabetes, the adoption of tools that allow early diagnosis and, lastly, the selection of appropriate treatment, i.e. the safest and most effective, supported by patient stratification models, in order to prevent/delay the development of diabetic complications

BRNS Sponsored National Symposium: Radiotracer Techniques: Improve Nutritional Status and Health of Animals and Plants

BRNS Sponsored National Symposium on "Radiotracer Techniques: Improve Nutritional Status and Health of Animals and Plants"  Organized By: Chameli Devi Institute of Pharmacy, Indore Saturday, 1st February 202

The emerging importance of lymphatics in health and disease: An NIH workshop report

The lymphatic system (LS) is composed of lymphoid organs and a network of vessels that transport interstitial fluid, antigens, lipids, cholesterol, immune cells, and other materials in the body. Abnormal development or malfunction of the LS has been shown to play a key role in the pathophysiology of many disease states. Thus, improved understanding of the anatomical and molecular characteristics of the LS may provide approaches for disease prevention or treatment. Recent advances harnessing single-cell technologies, clinical imaging, discovery of biomarkers, and computational tools have led to the development of strategies to study the LS. This Review summarizes the outcomes of the NIH workshop entitled Yet to be Charted: Lymphatic System in Health and Disease, held in September 2022, with emphasis on major areas for advancement. International experts showcased the current state of knowledge regarding the LS and highlighted remaining challenges and opportunities to advance the field

Current Medical Therapy and Revascularization in Peripheral Artery Disease of the Lower Limbs: Impacts on Subclinical Chronic Inflammation

Peripheral artery disease (PAD), coronary artery disease (CAD), and cerebrovascular disease (CeVD) are characterized by atherosclerosis and inflammation as their underlying mechanisms. This paper aims to conduct a literature review on pharmacotherapy for PAD, specifically focusing on how different drug classes target pro-inflammatory pathways. The goal is to enhance the choice of therapeutic plans by considering their impact on the chronic subclinical inflammation that is associated with PAD development and progression. We conducted a comprehensive review of currently published original articles, narratives, systematic reviews, and meta-analyses. The aim was to explore the relationship between PAD and inflammation and evaluate the influence of current pharmacological and nonpharmacological interventions on the underlying chronic subclinical inflammation. Our findings indicate that the existing treatments have added anti-inflammatory properties that can potentially delay or prevent PAD progression and improve outcomes, independent of their effects on traditional risk factors. Although inflammation-targeted therapy in PAD shows promising potential, its benefits have not been definitively proven yet. However, it is crucial not to overlook the pleiotropic properties of the currently available treatments, as they may provide valuable insights for therapeutic strategies. Further studies focusing on the anti-inflammatory and immunomodulatory effects of these treatments could enhance our understanding of the mechanisms contributing to the residual risk in PAD and pave the way for the development of novel therapies

Statische und dynamische Magnetfelder für die Nanopartikel-basierte zielgerichtete Wirkstofffreisetzung

Although medicine has made great progress in the last centuries and decades, it is still facing basic challenges that make doctors fail to efficiently and successfully treat the continuously emerging diseases and ailments due to ageing, industrialization, pollution and resulting biological mutations. In this context, the systemic chemotherapeutic treatment of cancer seems to be one of the most fitting examples for the wide gap between the usually followed medical approach and the theoretically optimal solution. Extrapolating from in vitro experiments and mouse models to humans, treating children as “miniaturized” adults when analyzing therapeutic effects, estimating drug doses based on relatively coarse processes like up scaling on weight, volume or area, and flooding the human body with drugs to solely achieve a minimal effect at the ailment site are just few examples for improvement needs in medical methods. One of the most promising approaches intended to bring more specificity and precision into the therapeutic toolbox is the directed delivery of drugs, already prophesized and described one hundred years ago by the German immunologist and Nobel Laureate in Medicine (1908) Paul Ehrlich (1854-1915) as the “magic bullet” principle. It is a visionary medical method in which active agents -such as drugs or antibodies- are guided within the human body and brought to bind directly and exclusively to their biological target. This approach was triggered and has been remarkably promoted by the introduction and continuous development of nano-sized medical systems since the 1950s, and is expected to experience a real breakthrough by the clinical validation of the so called “Magnetic Drug Targeting”. According to this technique, magnetically active nanoparticles are coated with a therapeutically active biomaterial and guided through external magnetic fields in the natural transport pathways of the body, then retained and concentrated at target sites where the biologically active load is set free. The delivered dose is augmented, side effects are lowered and the overall therapeutic efficiency is enhanced. Especially for cancer treatment, the magnetically guided drug delivery represents a huge potential. In fact, conventional chemotherapy methods are used systemically and succeed in best cases in delivering only a fractional amount of the drug to the target sites, while the rest is absorbed by the healthy tissue of the treated body. This is so inefficient that dose levels of about 50 to 100-fold those of conventional doses need to be administered to achieve cures of cancer cells (T. A. Connors 1995). As a result, blood filtering and trafficking organs, such as the liver, the kidneys, the spleen and most importantly the heart, are the direct victims of the highly toxic substances used in chemotherapy. Even the apparently more gentle approach of applying the maximum tolerated dose at defined intervals -in order to avoid toxicity- can unintentionally lead to a chemoresistance of the tumor (C. Damyanov 2009). These shortcomings of the chemical therapy further aggravate the fact that cancer is still the worldwide deadliest disease, with an upward trend. For instance, around 25 % of all registered death cases in the European Union are reported by the World Health Organization to be caused by tumors. Despite the development of advanced anti-cancer medicine, it still remains a difficult challenge to keep costs at an affordable level. For that reason, new and more efficient cancer treatment methods with higher success rates and lower side effects and costs are urgently needed and would help physicians cope with an ever ageing world population. In this work, we report improvements achieved in the understanding and control of the magnetically targeted drug delivery, mainly realized by the consideration of time issues and the investigation of dynamic magnetic fields. New approaches to assess the magnetic behavior of nanoparticles in suspensions as well as an advanced examination of the lung drug targeting and the mechanisms of cellular drug uptake after successful localized delivery represent the major achievements compiled in this manuscript. The registered improvements are an important contribution to the further development of the idea of directed therapies promoted by the emerging nanomedicine. This modern medicine is expected to provide techniques that can act on a cellular and even sub-cellular level, treating ailments with considerably more accuracy. Gradually, modern diagnostic and therapeutic techniques should elevate us slowly to the point where we can start thinking more in terms of real “regenerative” medicine. That means, we should be able to precisely and directly address pathologic tissues, save cells and organs, repair and heal them, rather than extinguish them.Mehr als hundert Jahre nach dem Tod von Paul Ehrlich, dem bedeutendsten deutschen Immunologen, verfolgt die "Nachwelt" noch mit großen Schritten eine seiner wichtigsten Visionen, die er während seiner Arbeiten zur Behandlung der Syphilis entwickelte: eine „Zauberkugel“ (magic bullet), die einen gegebenen krankmachenden Erreger gezielt abtöten kann. Ganz nach diesem noch -mehr denn je- aktuellen Prinzip, entwickeln Forscher heutzutage weltweit neue Methoden, um nicht nur Krankheitserreger, sondern auch befallene Gewebe, spezifisch zu behandeln. In den letzten Jahren entwickelte sich dadurch die Medizin von der konventionellen Anwendung, über die personalisierte Behandlung, wo die genetische Information eines jeden Patienten präventiv untersucht werden kann und die Ergebnisse zur Auswahl und Anpassung der Therapie-Art herangezogen werden, bis hin zur "Nanomedizin", einer neuen Ära der Arzneimittel-Konzipierung, -Synthese, -Dosierung und -Verabreichung, die Therapien auf zellulärer und sub-zellulärer Ebene ermöglichen sollte. Mediziner sind heutzutage weit entfernt von der Darstellung von Christian Friedrich Hebbel (18.03.1813 - 13.12.1863), dass "ein Arzt eine Aufgabe hat, als ob ein Mensch in einem dunklen Zimmer in einem Buche lesen sollte". Sie sind in der Lage, durch die Integration der Nanotechnologie im biomedizinischen Bereich, Gewebe und Zellen, die durchschnittliche Dimensionen von 10 µm haben, mit Nanosystemen im Submikrometer-Bereich zu adressieren und gezielt zu behandeln. In diesem Rahmen präsentiert sich das Magnetic Drug Targeting (MDT) als besonders wirksamer Therapie-Ansatz. Dabei werden Wirkstoff-beladene magnetische Nanopartikel über externe Magnetfelder im Körper geführt und an einem gegebenen Krankheitsort lokal angereichert. Die verabreichte Wirkdosis wird dadurch erhöht, Nebeneffekte minimiert. Besonders in der Krebsbekämpfung verspricht dieser Ansatz hohe Erfolgsquoten und eine Reduzierung der ohnehin enormen Chemo- und Radiotherapie-Kosten, die meistens einen bremsenden Effekt auf die Entwicklung und Verbreitung zahlreicher Behandlungsmethoden haben. An dieser Stelle sei daran erinnert, dass Krebs nach wie vor die weltweit wichtigste Todesursache ist, an der schätzungsweise 11.5 Millionen Weltbewohner im Jahre 2030 sterben werden, was einem Anstieg von 45% zum Jahre 2007 darstellt. Die zielgerichtete Arzneimittel-Applikation, zu Englisch "Directed Drug Delivery", soll hierfür Lösungen anbieten, die Tumore spezifisch angreifen und ausschalten können. Durch eine magnetische Lenkung und Anreicherung wird dieses Verfahren weiter optimiert. Die somit entstehende MDT-Methode eignet sich für Anwendungen in der Blutbahn, sowie in den Atemwegen von Patienten, mit entsprechenden Anpassungen. Entscheidend ist hierbei vor Allem das eingesetzte Magnetfeld, in Bezug auf Amplitude, Homogenität und Dynamik. In zahlreichen wissenschaftlichen Arbeiten, wurden bisher Erfolg versprechende Ergebnisse präsentiert, die überwiegend durch die Manipulation und Aufkonzentrierung von Nanopartikel-Wirkstoff-Komplexen mit statischen Magnetfeldern realisiert wurden. Eine hierzu komplementäre Betrachtung mit dynamischen Magnetfeldern wird in dieser Arbeit untersucht. Im Rahmen dieses Forschungsprojekts wurden Ansätze mit statischen und dynamischen Magnetfeldern zur Verbesserung des Magnetic Drug Targeting theoretisch überprüft, simulativ validiert und systemtechnisch umgesetzt. Nach einer ausführlichen Untersuchung der Nanopartikel-Eigenschaften, die den MDT-Effekt überhaupt ermöglichen und besonders beeinflussen, wurde der Anreicherungsprozess unter Magnetkraftwirkung modelliert und ein für Anwendungen in der Blutbahn optimiertes Magnetsystem simuliert, konstruiert und bei in-vivo-Versuchen eingesetzt. Dadurch konnte eine aktive und vor Allem reproduzierbare Retention von beladenen Nanopartikel-Komplexen in den Arterien und Venen der Rückenhaut einer Maus verzeichnet werden

Review on Buccal Adhesive Drug Delivery System: A Promising Strategy for Poorly Soluble Drugs

Rapid developments in the field of molecular biology and gene technology resulted in generation of many macromolecular drugs including peptides, proteins, polysaccharides and nucleic acids in great number possessing superior pharmacological efficacy with site specificity and devoid of untoward and toxic effects. However, the main impediment for the oral delivery of these drugs as potential therapeutic agents is their extensive presystemic metabolism, instability in acidic environment resulting into inadequate and erratic oral absorption. Parentral route of administration is the only established route that overcomes all these drawbacks associated with these orally less/inefficient drugs. But, these formulations are costly, have least patient compliance, require repeated administration, in addition to the other hazardous effects associated with this route. Over the last few decades' pharmaceutical scientists throughout the world are trying to explore transdermal and transmucosal routes as an alternative to injections. Among the various transmucosal sites available, mucosa of the buccal cavity was found to be the most convenient and easily accessible site for the delivery of therapeutic agents for both local and systemic delivery as retentive dosage forms, because it has expanse of smooth muscle which is relatively immobile, abundant vascularization, rapid recovery time after exposure to stress and the near absence of langerhans cells. Direct access to the systemic circulation through the internal jugular vein bypasses drugs from the hepatic first pass metabolism leading to high bioavailability. Further, these dosage forms are self-administrable, cheap and have superior patient compliance. Developing a dosage form with the optimum pharmacokinetics is a promising area for continued research as it is enormously important and intellectually challenging. With the right dosage form design, local environment of the mucosa can be controlled and manipulated in order to optimize the rate of drug dissolution and permeation. Advances in experimental and computational methodologies will be helpful in shortening the processing time from formulation design to clinical use

Effectiveness of intensive physiotherapy for gait improvement in stroke: systematic review

Introduction: Stroke is one of the leading causes of functional disability worldwide. Approximately 80% of post-stroke subjects have motor changes. Improvement of gait pattern is one of the main objectives of physiotherapists intervention in these cases. The real challenge in the recovery of gait after stroke is to understand how the remaining neural networks can be modified, to be able to provide response strategies that compensate for the function of the affected structures. There is evidence that intensive training, including physiotherapy, positively influences neuroplasticity, improving mobility, pattern and gait velocity in post-stroke recovery. Objectives: Review and analyze in a systematic way the experimental studies (RCT) that evaluate the effects of Intensive Physiotherapy on gait improvement in poststroke subjects. Methodology: Were only included all RCT performed in humans, without any specific age, that had a clinical diagnosis of stroke at any stage of evolution, with sensorimotor deficits and functional gait changes. The databases used were: Pubmed, PEDro (Physiotherapy Evidence Database) and CENTRAL (Cochrane Center Register of Controlled Trials). Results: After the application of the criteria, there were 4 final studies that were included in the systematic review. 3 of the studies obtained a score of 8 on the PEDro scale and 1 obtained a score of 4. The fact that there is clinical and methodological heterogeneity in the studies evaluated, supports the realization of the current systematic narrative review, without meta-analysis. Discussion: Although the results obtained in the 4 studies are promising, it is important to note that the significant improvements that have been found, should be carefully considered since pilot studies with small samples, such as these, are not designed to test differences between groups, in terms of the effectiveness of the intervention applied. Conclusion: Intensive Physiotherapy seems to be safe and applicable in post-stroke subjects and there are indications that it is effective in improving gait, namely speed, travelled distance and spatiotemporal parameters. However, there is a need to develop more RCTs with larger samples and that evaluate the longterm resultsN/

Gold nanoparticles for nanotheranostics in leukemia – Addressing Chronic Myeloid Leukemia

Leukemia is a type of cancer that initiates in the bone marrow and results in the unregulated production of immature white blood cells (leukemic cells). The most homogenous subgroup of the disease is chronic myeloid leukemia (CML) accounting for nearly 1.5 million patients worldwide. Virtually all cases harbor the genetic translocation t(9;22)(q34.1;q11.2) resulting in the BCR-ABL1 gene fusion, that encodes for BCR-ABL1 tyrosine kinase. CML treatment success relies on an early diagnosis and the intense research towards developing effective tyrosine kinase inhibitors (TKI). Nanotechnology offers unprecedent advantages to tackle the shortcomings of conventional procedures for the management of CML. Gold nanoparticles (AuNPs) have unique optical properties suitable for ex vivo biosensing applications, but can also function in vivo as nanocarriers in a theranostic approach that links treatment with diagnosis according to patient’s molecular profile. A gold nanoprobe (Au-nanoprobe) colorimetric assay was optimized for the detection of the most frequent BCR-ABL1 isoform (e14a2) and was validated on fully characterized clinical samples. This simple and cheap method enabled the direct detection of e14a2-expressing RNA samples, with accuracy and high specificity. The Au-nanoprobe assay was translated onto a microfluidics chip, resulting in a faster outcome with smaller sample volumes, due to the scale and design of the device. Additionally, a new therapeutic strategy was designed to overcome CML resistance to first line therapy, such as imatinib (IM). BCR-ABL1 gene silencing was effectively achieved in vitro, using AuNPs functionalized with polyethylene glycol and a hairpin-shaped antisense single stranded DNA (ssDNA) oligonucleotide. Furthermore, the nanoconstruct allowed to reduce the dose of IM, when tested in a combined approach, and potentiated a viability decrease of K562 cells resistant to IM. The results of this thesis strongly suggest that AuNPs are a suitable and flexible tool for CML nanotheranostics, improving detection and a personalized treatment strategy

The effects of celastrol on endothelial cells survival and proliferation

Introduction: Coronary artery bypass grafts are most commonly performed using saphenous vein grafts to complement the internal thoracic artery. The saphenous vein will remain popular despite its lower patency rate because it is easily accessible and lengthy enough to perform multiple bypasses. Therefore, several approaches have been studied, with the common goal of finding the optimal conditions that reduce graft failure. They include novel harvest techniques, new preservation preparations, innovative genetic therapies and experimental drugs. We believe a pharmacological pre-conditioning with an anti-oxidative and anti-inflammatory drug during the crucial time of harvest may spark beneficial survival response from the endothelial cells. One particular compound is Celastrol, an HSP90 inhibitor, which displays those antioxidant and anti-inflammatory properties. Methods: Human umbilical vein endothelial cells (HUVEC) were pretreated with various concentrations of Celastrol (10-10M, 10-8M and 10-6M). In order to reproduce oxidative stress found in ischemia/reperfusion, cells were exposed to hydrogen peroxide for a short and extended period (1h and 24h). To mimic storage condition encountered in clinical settings, cells were also exposed in heparinized normal saline. The viability was assessed by LIVE/DEAD assay. As for migrative and proliferative properties, scratch tests were performed. Finally, various protective intracellular pathways were evaluated by Western blot. Results: This study shows that pre-treatment with Celastrol promotes survival in HUVEC submitted to oxidative stress. Notable improvement in cellular viability was detected as early as 1 hour after oxidative stress (H2O2 4 mM), 76.6% vs 66.1% (p=0.005). Significant survival benefits are also reported after prolonged oxidative stress (H2O2 0.5 mM for 24 hours); viability was 93.7% vs 76.9% (p=0.001) for Cel 10- 8 M and 96.6% vs 76.9% (p=0.002) for Celastrol 10-10M when compared to the vehicle. Celastrol, however, did not significantly affect viability of HUVEC stored in heparinized normal saline. Celastrol at 10-6 M promotes faster and more complete wound closure compared to the vehicle or to lower dosages. Celastrol triggers early activation of the RISK pathway, inducing activation of both Akt and ERK1/2 within the first 15 minutes of treatment. Celastrol also induces the expression of HSP70 and HO-1, effectors of the Heat Shock Response and the anti-oxidative response respectively. Conclusion: Pre-treatment by Celastrol provides survival benefits in endothelial cells under oxidative stress. It also stimulates endothelial cell proliferation and migration, promoting faster and more complete re-endothelialisation. Celastrol can potentially be used as an additive to storage solutions to limit endothelial injury and promote graft protection.Introduction: La chirurgie de pontage coronarien requiert, dans la grande majorité des cas, l’utilisation de l’artère mammaire interne en combinaison avec un ou des greffons provenant de la grande veine saphène. Malgré le taux de perméabilité inférieur aux artères, la veine saphène reste un choix populaire de conduit en raison de son accessibilité et de sa longueur. De ce fait, le greffon veineux devient la cible de multiples approches et le sujet de nombreuses études visant à optimiser sa perméabilité. Celles-ci incluent le raffinement des techniques de prélèvement, les solutions de préservations, les agents pharmacologiques ainsi que la thérapie génique. Il est davantage intéressant de combiner les approches afin de joindre leurs bénéfices, comme, par exemple, ajouter un agent pharmacologique à une solution de préservation. Un agent potentiel serait le Celastrol, connu pour être un inhibiteur du HSP90 et possède des propriétés antioxydantes et anti-inflammatoires. Méthodologie: Des cellules endothéliales humaines provenant de la veine ombilicale (HUVEC) sont pré-conditionnées à de multiples concentrations de Celastrol (10-10M, 10-8M and 10-6M) pendant une heure avant d’être soumises aux conditions de stress. Pour reproduire les conditions per-opératoires de prélèvement, les cellules endothéliales ont été préservées dans du salin (NS) héparinisé. Pour mimer le stress secondaire à l’ischémie/reperfusion, les cellules ont aussi été soumises à diverses concentrations de H2O2. Une analyse de la viabilité cellulaire fut conduite par le test de LIVE/DEAD. La capacité de ré-endothélialisation est étudiée grâce à l’épreuve de scratch test. Les voies intracellulaires de survie telles que le RISK pathway (Akt, ERK1/2), le Heat shock response (HSP70) et la réponse anti-oxydante (via l’activité de HO-1) ont été examinées par immunoblot. Résultats: Les résultats démontrent que la préservation des cellules endothéliales dans du NS héparinisé est associée à une augmentation de la mortalité comparativement au milieu de culture (20.4% vs 1.9%, p=0.004). Toutefois, un traitement au Celastrol n’affecte pas significativement la survie des cellules endothéliales dans le NS héparinisé. Le stress oxydatif induit aussi une augmentation de la mortalité, et ce à dose-dépendante. Suivant un court stress 6 oxydatif (H2O2 4 mM), un pré-traitement au Celastrol 10-10M est associé à une meilleure viabilité comparativement au véhicule (76.6% vs 66.1%, p=0.005). Lorsque soumises à un stress oxydatif prolongé (H2O2 0.5 mM pendant 24h), les HUVEC pré-traitées au Celastrol à 10-8M et 10-10M démontrent une amélioration significative de la viabilité, 93.7% vs 76.9% (p=0.001) et 93.6% vs 76.9% (p=0.002) respectivement. Quant à la ré-endothélialisation, un traitement au Celastrol 10- 6M est associé à une fermeture plus rapide et complète comparativement au véhicule. Un court traitement au Celastrol active précocement les kinases de la voie de RISK (Akt et ERK). Le traitement induit aussi l’expression de HSP70 et HO-1 qui reste soutenue jusqu’à 48 heures posttraitement. Conclusion: Le Celastrol active plusieurs voies de protection intracellulaire tels que le RISK pathway, le Heat Shock Response et la réponse antioxydante via l’activité de HO-1. En corrélation avec cette réponse, il améliore la survie des cellules endothéliales dans un milieu oxydatif. Le Celastrol promeut aussi une ré-endothélialisation plus complète et rapide. Cette étude met en valeur les bénéfices potentiels du Celastrol sur les cellules endothéliales. Afin d’optimiser la protection du greffon, le Celastrol pourrait donc être considéré comme agent adjuvant à une solution de préservation