Estabelecimento e caracterização de modelos resistentes e matastaticos de osteossarcoma humano in vitro e in vivo

Doutoramento em BiologiaOsteossarcoma é uma doença rara, sendo o tipo mais comum de tumor maligno do osso. O pico de incidência ocorre durante a adolescência e desenvolve-se principalmente nos ossos longos. Os tratamentos atuais incluem quimioterapia antes e após a cirurgia e a ressecção cirúrgica de todos os locais envolvidos (tumor primário e metástases quando presente). As metástases, principalmente nos pulmões, são um grande problema no diagnóstico (20-30% dos pacientes) e durante a história natural do osteossarcoma (cerca de 30% de recaída), afetam uma percentagem considerável de pacientes e são considerados os maiores problemas desta doença. Biologicamente, os osteossarcomas são um dos tumores mais complexos observados nas crianças, no que diz respeito à heterogeneidade, anomalias moleculares e cromossómicas e ao seu microambiente específico. A resistência aos agentes quimioterápicos utilizados no tratamento do osteossarcoma também é um fator prognóstico de alto risco de recaída, independentemente da quimioterapia utilizada. É urgente compreender os mecanismos relacionados a esses fenómenos e desenvolver novos quimioterápicos para superar esses problemas e aumentar a taxa de sobrevivência do paciente. O desenvolvimento de novos fármacos requer múltiplos modelos pré-clínicos adequados para mimetizar a complexidade genómica do osteossarcoma que se desenvolve-se num microambiente ósseo e metastático nos pulmões, apesar dos tratamentos quimioterápicos habituais. Nesta tese, foram desenvolvidos e caracterizados diferentes modelos pré-clínicos clinicamente relevantes in vitro e in vivo, incluindo modelos resistentes bioluminescentes, de modo a melhor compreender esta doença e alguns dos mecanismos de resistência relacionados. Desenvolvemos, em primeiro lugar, dois modelos ortotópicos xenotransplantados derivados de linhas celulares (CDX) bioluminescentes (Luc/mKate2), capazes de desenvolver metástases espontaneamente. As células bioluminescentes foram injetadas ortotopicamente, em diferentes contextos: imune (estirpes de ratinhos de laboratório - nude e NSG) e ósseo (intratibial e paratibial com ativação do periósteo). O sistema IVIS SpectrumCT, combinando tomografia computadorizada longitudinal (TC) e bioluminescência, foi utilizado para acompanhar o crescimento primário do tumor e a disseminação metastática em tempo real. O contexto imune murino, o contexto genético dos dois modelos CDX e o contexto ósseo (intratibial ou paratibial) influenciaram o enxerto tumoral, o crescimento primário do tumor e o comportamento agressivo local (osteocondensação e osteólise), bem como a disseminação metastática para os pulmões, ossos e baço (uma localização incomum em seres humanos). Observou-se também que a estirpe de ratinhos NSG e a injeção intratibial apresentam melhores características para o desenvolvimento de modelos que a injeção paratibial ou a estirpe de ratinhos nude. Seguidamente, desenvolvemos modelos resistentes bioluminescentes in vitro, aos principais medicamentos utilizados no osteossarcoma, nomeadamente metotrexato (5modelos) e doxorrubicina (1modelo), por exposição contínua a esses medicamentos. Realizando o mesmo procedimento, não foi obtida resistência à mafosfamida. Investigamos os mecanismos da resistência adquirida relacionados com estas drogas e observamos comportamentos diferenciais in vitro e in vivo (com modelos CDX ortotópicos bioluminescentes) das linhas resistentes e respetivas linhas parentais. Um mecanismo de resistência na linha celular resistente à doxorrubicina foi observado, nomeadamente a indução da proteína PgP. Mostramos diferentes mecanismos de resistência adquirida ao metotrexato de acordo com o backgroud genético das linhas celulares, que afetam a expressão génica e provocam alterações no número de cópias ao nível dos cromossomas. Foram observados diferentes comportamentos dos modelos resistentes bioluminescentes ortotópicos (CDX) in vivo em comparação com as respetivas linhas parentais. Finalmente, utilizando amostras de osteossarcoma humano provenientes de biópsias de pacientes em recidiva após a quimioterapia habitual, foram desenvolvidos modelos resistentes xenotransplantados derivados do paciente (PDX), quer subcutaneamente quer ortotopicamente (no osso). A caracterização desses modelos está em curso, em particular a comparação das características moleculares destes (sequenciamento completo do exoma e sequenciamento do ARN) com as do tumor do paciente na recaída e do mesmo no diagnóstico. Todos esses modelos desenvolvidos em diferentes contextos in vitro e in vivo trazem informações complementares para outros tipos de modelos de osteossarcoma já existentes. Estes modelos são necessários para obter mais informações sobre os diferentes processos que envolvem o desenvolvimento inicial, a progressão e a sensibilidade/resistência ao tratamento no osteossarcoma. Permitem ajudar ainda a avaliação de novos quimioterápicos, de modo a encontrar soluções para a atual falta de terapias eficientes no osteossarcoma.Osteosarcoma is a rare disease and the most common type of malignant bone tumor. The peak incidence occurs during the adolescence and the disease develops mainly in long bones. Current treatments include chemotherapy before and after surgery and surgical resection of all the involved sites (primary tumor and metastasis when present). Metastases mainly in the lungs are a major challenge at diagnosis (20-30% of the patients) and during the natural history of osteosarcoma (around 30% of relapse, most being metastatic), affect a considerable percentage of patients with osteosarcoma, being considered the biggest problem of this disease. Biologically, osteosarcomas are one of the most complex tumours in children in regard to tumour heterogeneity, molecular and chromosomal abnormalities, and their specific microenvironment. Resistance to the chemotherapeutic agents used in osteosarcoma is also a prognostic factor of high risk of relapse, whatever the chemotherapy used. It is urgent to understand the mechanisms related with these phenomena and develop new drugs in order to overcome these challenges and increase patient survival. New drug development requires suitable multiple pre-clinical models to better mimic the genomic complexity of osteosarcoma which develops in a bone microenvironment and in a metastatic setting in the lungs, despite usual chemotherapeutic treatments. In this thesis, we developed and characterised different and clinically relevant in vitro and in vivo preclinical models, including bioluminescent resistant models in order to understand better this disease and some of the resistant mechanism related. First, two bioluminescent (Luc/mKate2) cell line derived xenograft (CDX) models were developed in an orthotopic bone setting able to spontaneously metastasize. Bioluminescent cells were injected orthotopically, in different immune (nude and NSG mouse strains) and bone (intratibial and paratibial with periosteum activation) contexts. IVIS SpectrumCT system, combining longitudinal computed tomography (CT) and bioluminescence, was used to follow primary tumor growth and metastatic spread in real-time. The murine immune context, the genetic background of the two CDX-models, and the bone context (intratibial or paratibial) influenced tumor engraftment, primary tumor growth and local aggressive behavior (osteocondensation and osteolysis) as well as metastatic spread to lungs, bone, and spleen (an unusual localization in humans). It was also observed that intratibial injection in NSG mice showed better characteristics for model development than paratibial injection or nude mice recipient. We further developed in vitro bioluminescent models that were resistant to the main drugs used in osteosarcoma, methotrexate (5 models) and doxorubicin (one model), by continuous exposure to these drugs. With the same technique no resistance was obtained for mafosfamide. We explored the mechanism of the acquired resistance to these drugs and observed the differential in vitro and in vivo behaviors (with CDX bioluminescent orthotopic models) of the resistant lines and their parental counterpart. A multidrug phenomenon by PgP induction was observed in the doxorubicin resistant cells. We show different mechanisms of acquired resistance to methotrexate according to the genetic background of the cell lines affecting either gene expression and copy number abnormalities. Different in vivo behavior of the resistant bioluminescent orthotopic CDX models compared to their parental counterparts were observed. Finally, using human biopsy samples of osteosarcoma relapsing after usual anti-osteosarcoma chemotherapy were developed resistant patient-derived xenograft (PDX) models, either in subcutaneous as in orthotopic bone setting. The characterization of these models are still ongoing, in particular the comparison of their molecular characteristics, i.e. using whole exome and RNA sequencing, in comparison with the patient tumor at relapse and with the same patient tumor at diagnosis. All these multiple models developed in different in vitro and in vivo contexts bring complementary information to other types of existing osteosarcoma models. They are needed to get more insight into the different processes involving osteosarcoma initiation, progression and in particular treatment sensitivity/resistance. They will further help drug testing to find solution to the current lack of efficient new drugs in osteosarcoma

Indução de stress oxidativo pelo sulfurafano em osteosarcoma humano

Mestrado em Biologia Aplicada - Microbiologia Clínica e AmbientalO sulforafano (SFN) é um composto que pertence à família dos isotiocianatos, sendo o mais estudado deste grupo, principalmente pelas suas potenciais propriedades anti-tumorais. Existe em grande quantidade nas crucíferas, sobretudo nos brócolos. Vários estudos têm demonstrado a capacidade do SFN em causar paragem do ciclo celular, indução da apoptose, aumento de espécies reactivas de oxigénio (ROS), no entanto, os seus efeitos no osteosarcoma, estão ainda pouco compreendidos, principalmente ao nível do stress oxidativo. O objectivo deste estudo foi investigar a capacidade do SFN induzir, in vitro, stress oxidativo numa linha celular de osteossarcoma humano, a MG-63, e compreender as vias de stress oxidativo envolvidas neste processo. Para tal, as células MG-63 foram expostas a diferentes concentrações de SFN (0, 5, 10 e 20μM) em diferentes períodos (24 e 48h). O crescimento e a morfologia celular foram observados através do microscópio invertido e a viabilidade celular determinada usando o ensaio de Trypan Blue. A avaliação do stress oxidativo foi realizada através da análise da actividade das enzimas envolvidas na destoxificação de espécies reativas de oxigénio (ROS), nomeadamente glutationa peroxidase, glutationa reductase e superóxido dismutase. A actividade antioxidante total, a glutationa reduzida e a peroxidação lipídica foram também analisadas, bem como a expressão de genes relacionados com o stress oxidativo (SOD1, SOD2, GSR, GPX1, GSTM1 e GSTM4 e CAT). Os ensaios enzimáticos, bem como a actividade antioxidante total, foram avaliadas por diferentes ensaios colorimétricos no leitor de microplacas, já a glutationa reduzida e a peroxidação lipídica foram avaliadas por fluorimetria. Observou-se para ambos os períodos de exposição que a maior concentração de SFN reduziu a viabilidade celular, relativamente ao controlo, bem como a actividade de todas as enzimas analisadas. A actividade antioxidante total aumenta com o tempo e diminui com a concentração de SFN. A actividade das enzimas decresce com o aumento da concentração de SFN (e com o tempo para concentrações de SFN elevadas). Verificou-se também um aumento da glutationa reduzida para concentrações de SFN baixas, embora tenha decrescido para valores de SFN mais elevadas. A expressão dos genes é concordante em geral com a actividade das respectivas enzimas, ou seja diminui com o aumento da concentração. No entanto é observada uma excepção para a SOD em que a actividade diminui com a concentração enquanto a expressão aumenta. Pode-se observar que a peroxidação lipídica diminui com a concentração. Como conclusão, estes resultados indicam que o SFN induz stress oxidativo em células de osteossarcoma humano. Propõe-se um modelo de acção para o stress oxidativo induzido por SFN em osteossarcoma.Sulforaphane (SFN) is a compound that belongs to the isothiocyanates family and is the most studied compound of this group, mostly due to its putative anti-tumoral properties. It is found in large quantities in cruciferous vegetables, mainly broccoli. Several studies have shown the ability of SFN to cause cell cycle arrest, apoptosis induction, reactive oxygen species (ROS) formation however, its effects in osteosarcoma, is still poorly understood, mostly at the stress oxidative level. The aim of this study was to investigate the ability of SFN in inducing, in vitro, oxidative stress in the human osteosarcoma cell line MG-63. It was also an aim to understand the pathways of oxidative stress involved in this process. For these propose, MG-63 cells were exposed to different concentrations of SFN (0, 5, 10 and 20μM) at different times (24 and 48h). The growth and cell morphology were observed through an inverted microscope, and cell viability determined using Trypan Blue assay. The oxidative stress evaluation was performed by the activity analysis of antioxidant enzymes, namely, glutathione peroxidase, superoxide dismutase and glutathione reductase. The total antioxidant activity (TAA), reduced glutathione and lipid peroxidation were also analysed, as well as the expression of genes related to oxidative stress (SOD1, SOD2, GSR, GPX1, GSTM1 and GSTM4 and CAT). Enzyme assays and total antioxidant activity were evaluated by different colorimetric assays on a microplate reader, while the reduced glutathione and lipid peroxidation were evaluated by fluorimetry. It was observed (for both exposure periods) that the highest concentration of SFN reduced cell viability relatively to control, and reduced the activity of all tested enzymes. The total antioxidant activity increased with time and decreased with the concentration of SFN. The activity of enzymes decreased with the increasing concentration of SFN (and with time for the highest SFN concentrations). There was also an increase of the reduced glutathione for low SFN concentrations, while it decreased for higher concentrations of SFN. Overall, the gene expression is consistent with the corresponding enzymes activities, i.e., the expression decreased with increasing concentrations of SFN. However, SOD was an exception (SOD activity decreased with SFN concentration whereas its expression increased). It can be observed that lipid peroxidation declined with SFN concentration increase. Finally, these results indicate that SFN induces oxidative stress in human osteosarcoma cells. It is proposed a model for the pathways involved in this oxidative stress–induced by SFN in osteosarcoma

A biobank of pediatric patient-derived-xenograft models in cancer precision medicine trial MAPPYACTS for relapsed and refractory tumors

Cancer models; Paediatric cancerModelos de cáncer; Cáncer pediátricoModels de càncer; Càncer pediàtricPediatric patients with recurrent and refractory cancers are in most need for new treatments. This study developed patient-derived-xenograft (PDX) models within the European MAPPYACTS cancer precision medicine trial (NCT02613962). To date, 131 PDX models were established following heterotopical and/or orthotopical implantation in immunocompromised mice: 76 sarcomas, 25 other solid tumors, 12 central nervous system tumors, 15 acute leukemias, and 3 lymphomas. PDX establishment rate was 43%. Histology, whole exome and RNA sequencing revealed a high concordance with the primary patient’s tumor profile, human leukocyte-antigen characteristics and specific metabolic pathway signatures. A detailed patient molecular characterization, including specific mutations prioritized in the clinical molecular tumor boards are provided. Ninety models were shared with the IMI2 ITCC Pediatric Preclinical Proof-of-concept Platform (IMI2 ITCC-P4) for further exploitation. This PDX biobank of unique recurrent childhood cancers provides an essential support for basic and translational research and treatments development in advanced pediatric malignancies.This work was supported by grants from Fondation Gustave Roussy; Fédération Enfants Cancers et Santé, Société Française de lutte contre les Cancers et les leucémies de l’Enfant et l’adolescent (SFCE), Association AREMIG and Thibault BRIET; Parrainage médecin-chercheur of Gustave Roussy; INSERM; Canceropôle Ile-de-France; Ligue Nationale Contre le Cancer (Equipe labellisée); Fondation ARC for the European projects ERA-NET on Translational Cancer Research (TRANSCAN 2) Joint Transnational Call 2014 (JTC 2014) ‘Targeting Of Resistance in PEDiatric Oncology (TORPEDO)’, ERA-NET TRANSCAN JTC 2014 (TRAN201501238), and TRANSCAN JTC 2017 (TRANS201801292); Agence Nationale de la Recherche (ANR-10-EQPX-03, Institut Curie Génomique d’Excellence (ICGex); IMI ITCC-P4; The Child Cancer Research Foundation (CCRF), Cancer Council Western Australia (CCWA); PAIR-Pédiatrie/CONECT-AML (INCa-ARC-LIGUE_11905 and Association Laurette Fugain), Ligue contre le cancer (Equipe labellisée, since 2016), OPALE Carnot institute; Dell; Fondation Bristol-Myers Squibb; Association Imagine for Margo; Association Manon Hope; L’Etoile de Martin; La Course de l’Espoir; M la vie avec Lisa; ADAM; Couleur Jade; Dans les pas du Géant; Courir pour Mathieu; Marabout de Ficelle; Olivier Chape; Les Bagouz à Manon; Association Hubert Gouin Enfance et Cancer; Les Amis de Claire; Kurt-und Senta Hermann Stiftung; Holcim Stiftung Wissen; Gertrud-Hagmann-Stiftung für Malignom-Forschung; Heidi Ras Grant Forschungszentrum fürs Kind; Children’s Liver Tumor European Research Network (ChiLTERN) EU H2020 projet (668596); Fundación FERO and the Rotary Clubs Barcelona Eixample, Barcelona Diagonal, Santa Coloma de Gramanet, München-Blutenburg, Sassella-Stiftung, Berger-Janser Stiftung and Krebsliga Zürich, Deutschland Gemeindienst e.V. and others from Barcelona and province, and No Limits Contra el Cáncer Infantil Association

Combination Therapies Targeting ALK-aberrant Neuroblastoma in Preclinical Models

PURPOSE ALK-activating mutations are identified in approximately 10% of newly diagnosed neuroblastomas and ALK amplifications in a further 1%-2% of cases. Lorlatinib, a third-generation anaplastic lymphoma kinase (ALK) inhibitor, will soon be given alongside induction chemotherapy for children with ALK-aberrant neuroblastoma. However, resistance to single-agent treatment has been reported and therapies that improve the response duration are urgently required. We studied the preclinical combination of lorlatinib with chemotherapy, or with the MDM2 inhibitor, idasanutlin, as recent data have suggested that ALK inhibitor resistance can be overcome through activation of the p53-MDM2 pathway. EXPERIMENTAL DESIGN We compared different ALK inhibitors in preclinical models prior to evaluating lorlatinib in combination with chemotherapy or idasanutlin. We developed a triple chemotherapy (CAV: cyclophosphamide, doxorubicin, and vincristine) in vivo dosing schedule and applied this to both neuroblastoma genetically engineered mouse models (GEMM) and patient-derived xenografts (PDX). RESULTS Lorlatinib in combination with chemotherapy was synergistic in immunocompetent neuroblastoma GEMM. Significant growth inhibition in response to lorlatinib was only observed in the ALK-amplified PDX model with high ALK expression. In this PDX, lorlatinib combined with idasanutlin resulted in complete tumor regression and significantly delayed tumor regrowth. CONCLUSIONS In our preclinical neuroblastoma models, high ALK expression was associated with lorlatinib response alone or in combination with either chemotherapy or idasanutlin. The synergy between MDM2 and ALK inhibition warrants further evaluation of this combination as a potential clinical approach for children with neuroblastoma

Recent advances in understanding osteosarcoma and emerging therapies

International audienceOsteosarcoma is the most common bone cancer in adolescents and young adults, but it is a rare cancer with no improvement in patient survival in the last four decades. The main problem of this bone tumor is its evolution toward lung metastatic disease, despite the current treatment strategy (chemotherapy and surgery). To further improve survival, there is a strong need for new therapies that control osteosarcoma cells with metastatic potential and their favoring tumor microenvironment (ME) from the diagnosis. However, the complexity and heterogeneity of those tumor cell genomic/epigenetic and biology, the diversity of tumor ME where it develops, the sparsity of appropriate preclinical models, and the heterogeneity of therapeutic trials have rendered the task difficult. No tumor- or ME-targeted drugs are routinely available in front-line treatment. This article presents up-to-date information from preclinical and clinical studies that were recently published or presented in recent meetings which we hope might help change the osteosarcoma treatment landscape and patient survival in the near future

MT2A is an early predictive biomarker of response to chemotherapy and a potential therapeutic target in osteosarcoma

International audienceOsteosarcoma is the most prevalent primary bone malignancy in children and young adults. Resistance to chemotherapy remains a key challenge for effective treatment of patients with osteosarcoma. The aim of the present study was to investigate the preventive role of metallothionein-2A (MT2A) in response to cytotoxic effects of chemotherapy. A panel of human and murine osteosarcoma cell lines, modified for MT2A were evaluated for cell viability, and motility (wound healing assay). Cell-derived xenograft models were established in mice. FFPE tumour samples were assessed by IHC. In vitro experiments indicated a positive correlation between half-maximal inhibitory concentration (IC50) for drugs in clinical practice, and MT2A mRNA level. This reinforced our previously reported correlation between MT2A mRNA level in tumour samples at diagnosis and overall survival in patients with osteosarcoma. In addition, MT2A/MT2 silencing using shRNA strategy led to a marked reduction of IC50 values and to enhanced cytotoxic effect of chemotherapy on primary tumour. Our results show that MT2A level could be used as a predictive biomarker of resistance to chemotherapy, and provide with preclinical rational for MT2A targeting as a therapeutic strategy for enhancing anti-tumour treatment of innate chemo-resistant osteosarcoma cells

In-Vitro and In-Vivo Establishment and Characterization of Bioluminescent Orthotopic Chemotherapy-Resistant Human Osteosarcoma Models in NSG Mice

International audienceOsteosarcoma, the most common bone malignancy with a peak incidence at adolescence, had no survival improvement since decades. Persistent problems are chemo-resistance and metastatic spread. We developed in-vitro osteosarcoma models resistant to chemotherapy and in-vivo bioluminescent orthotopic cell-derived-xenografts (CDX). Continuous increasing drug concentration cultures in-vitro resulted in five methotrexate (MTX)-resistant and one doxorubicin (DOXO)-resistant cell lines. Resistance persisted after drug removal except for MG-63. Different resistance mechanisms were identified, affecting drug transport and action mechanisms specific to methotrexate (RFC/SCL19A1 decrease, DHFR up-regulation) for MTX-resistant lines, or a multi-drug phenomenon (PgP up-regulation) for HOS-R/DOXO. Differential analysis of copy number abnormalities (aCGH) and gene expression (RNAseq) revealed changes of several chromosomic regions translated at transcriptomic level depending on drug and cell line, as well as different pathways implicated in invasive and metastatic potential (e.g., Fas, Metalloproteinases) and immunity (enrichment in HLA cluster genes in 6p21.3) in HOS-R/DOXO. Resistant-CDX models (HOS-R/MTX, HOS-R/DOXO and Saos-2-B-R/MTX) injected intratibially into NSG mice behaved as their parental counterpart at primary tumor site; however, they exhibited a slower growth rate and lower metastatic spread, although they retained resistance and CGH main characteristics without drug pressure. These models represent valuable tools to explore resistance mechanisms and new therapies in osteosarcoma

A biobank of pediatric patient-derived-xenograft models in cancer precision medicine trial MAPPYACTS for relapsed and refractory tumors

Abstract Pediatric patients with recurrent and refractory cancers are in most need for new treatments. This study developed patient-derived-xenograft (PDX) models within the European MAPPYACTS cancer precision medicine trial (NCT02613962). To date, 131 PDX models were established following heterotopical and/or orthotopical implantation in immunocompromised mice: 76 sarcomas, 25 other solid tumors, 12 central nervous system tumors, 15 acute leukemias, and 3 lymphomas. PDX establishment rate was 43%. Histology, whole exome and RNA sequencing revealed a high concordance with the primary patient’s tumor profile, human leukocyte-antigen characteristics and specific metabolic pathway signatures. A detailed patient molecular characterization, including specific mutations prioritized in the clinical molecular tumor boards are provided. Ninety models were shared with the IMI2 ITCC Pediatric Preclinical Proof-of-concept Platform (IMI2 ITCC-P4) for further exploitation. This PDX biobank of unique recurrent childhood cancers provides an essential support for basic and translational research and treatments development in advanced pediatric malignancies

Ciência, Crise e Mudança. 3.º Encontro Nacional de História das Ciências e da Tecnologia. ENHCT2012

III Encontro Nacional de História das Ciências e da Tecnologia. O Centro de Estudos de História e Filosofia da Ciência, organiza o 3.º Encontro Nacional de História da Ciência e da Técnica, sob o tema «Ciência, Crise e Mudança» que tem lugar na Universidade de Évora, nos dias 26, 27 e 28 de Setembro de 2012. O Primeiro Encontro Nacional de História da Ciência teve lugar em 21 e 22 Julho de 2009, no seguimento do programa de estímulo ao de¬senvolvimento da História da Ciência em Portugal e de valorização do património cultural e científico do País, lançado pelo Ministério da Ciência, Tecnologia e Ensino Superior (MCTES) em 31 de Janeiro desse ano. A sua organização coube a investigadores do Instituto de História Contemporânea (IHC), da FCSH da UNL, e do Centro Científico e Cultural de Macau (CCCM), em cujas instalações se realizou. De en¬tre as conclusões do Encontro, destacou-se a de realizar periodicamen¬te novos Encontros Nacionais, a serem organizados de forma rotativa por diferentes centros e núcleos de investigadores. Na sequência deste Primeiro Encontro, o Centro Interuniversitário de História das Ciências e da Tecnologia (CIUHCT) organizou, entre 26 e 28 de Julho de 2010, o II Encontro, dedicado ao tema “Comunicação das Ciências e da Tecnologia em Portugal: Agentes, Meios e Audiências”. Cabe agora ao CEHFCi cumprir o que foi decidido no final deste Encontro. Na situação económica e política que hoje vivemos torna-se particularmente urgente aprofundar o estudo e o debate sobre a interação entre a Sociedade, a Ciência e a sua História. Coordenação Científica e Executiva do encontro estiveram a cargo de dois investigadores CEHFCi: Maria de Fátima Nunes, José Pedro Sousa Dia