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Agradecimentos: This work was supported by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Abstract: Chronic graft-versus-host disease (cGvHD), an immunological complication of allogeneic cell transplantation, is the principal cause of non-relapse mortality and morbidity. Even though advances have been made in understanding the pathophysiology of this disorder, many questions remain. We sought to evaluate gene expression of transforming growth factor ß (TGF-ß) pathway components, through quantitative RT-PCR and PCR array, in patients with cGvHD with different disease activity. We observed an upregulation of SMAD3, BMP2, CDKN1A, IL6, and TGF-ß2 genes in the clinical tolerance group, which had never developed cGvHD, or which had been withdrawn from all immunosuppressive treatments (IST) for at least 1 year. In addition, SMAD5 gene upregulation was observed in cGvHD patients undergoing IST, and ordinal regression showed a correlation between SMAD5 expression and disease severity. Our data support the evidence of the important role of TGF-ß effects in the pathological process of cGvHD FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO (FAPESP) CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO (CNPQ) Fechado

Agradecimentos: The authors would like to thank Raquel S Foglio (Hematology and ransfusion Medicine Center, University of Campinas, São Paulo, Brazil) for the English revision; Ana Leda Longhini (University of Alabama, Birmingham, AL, USA) and Irene Santos (Hematology and Transfusion Medicine Center, University of Campinas, São Paulo, Brazil) for ow cytometry assistance; Renata Giardini Rosa and Juares Ednaldo Romero Bianco for the evelopment of scaffold material; and Alessio Molinari and Maurizzio Bottas (in memoriam) Dipartimento di Biotecnologie, Chimica e Farmacia-Università degli Studi di Siena, Siena, Italy) or the synthesis of iHCK-37 compound. We also thank the National Institute of Science and echnology on Photonics Applied to Cell Biology (INFABIC) at UNICAMP for the access to quipment and confocal assistance and analysis (FAPESP Grants 2014/50938-8 and CNPq Grants 465699/2014-6) and the Hemostasis Laboratory of Hematology and Transfusion Medicine enter (FAPESP Grants 2016/14172-6) at UNICAMP for cell proliferation analysis by IncuCyte S3 system (Sartorius). The authors also thank CNPq, FAPESP, and INCT-Sangue for the fnancial support Abstract: Myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are characterized by risk of relapses, poor survival, unwanted side efects and high toxicity with the current therapies. In light of these facts, there are eforts to develop new drugs specifc for deregulated molecules that participate in leukemia pathogenesis. Hematopoietic cell kinase (HCK), an Src kinase family member, is overexpressed on hematopoietic stem cells of MDS and de novo AML patients and involved in the oncogenic process. Thus, we investigated in vitro, ex vivo and in vivo efects of a novel chemical compound targeting HCK inhibition (iHCK-37), in combination with the most used drugs for the treatment of MDS and de novo AML, 5-Azacytidine and Cytarabine. Herein, the combination treatment with iHCK-37 and 5-Azacytidine or Cytarabine demonstrated additive efects against leukemia cells, compared to either drug alone. iHCK-37 plus 5-Azacytidine or Cytarabine treatment was able to reduce the activation of oncogenic pathways, MAPK/ERK and PI3K/AKT, leading to reduction of ERK and AKT phosphorylation, and increased BAX and decreased BCL-XL protein expression. Moreover, treatment with iHCK-37 reduced MDS and AML CD34-positive cell numbers inside a 3D-structure but did not afect normal CD34-positive cell numbers. In vivo analysis showed that leukemic mice treated with iHCK-37 had reduced ERK and AKT proteins phosphorylation levels and leukocyte numbers. In conclusion, the iHCK-37 inhibitor has anti-neoplastic activity in leukemia cells without altering apoptosis and survival rate of normal cells, suggesting on-target malignant cell killing activity as a single agent or in combination with 5-Azacytidine or Cytarabine FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQ Fechado

Orientador : Marco A. Zago Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas Resumo: A etiopatogenia das úlceras de perna nas doenças falciformes é, até o momento, desconhecida, ainda que a vasooclusão possa ser reconhecida como fator causal. Com o objetivo de avaliar o possível papel de obstrução ao fluxo circulatório na gênese das úlceras de perna na anemia falciforme, foi empregado o método de medida da depuração de Tc99m, injetado no tecido muscular, no repouso e após exercício. Neste estudo, foram demonstradas diferenças na velocidade de eluição de Tc99m injetado na musculatura gastrocnêmica de pacientes com anemia falciforme quando comparados a indivíduos sem anemia e obstrução arterial ou pacientes com anemias adquiridas, indicando a ocorrência de anormalidades do fluxo circulatório periférico. O estudo incluiu 27 pacientes com anemia falciforme em fase estável (88), dos quais 13 nunca haviam apresentado úlceras de perna, 20 controles sem anemia e doença vascular (N) e 20 pacientes com anemias adquiridas (AA) .Para padronização do método foram estudados 20 pacientes com doença vascular obstrutiva periférica (C I>. No repouso, os dois grupos de indivíduos anêmicos apresentaram velocidade de eluição de Tc99m superior ao grupo N. No entanto, esta diferença só foi significativa para aqueles com anemia falciforme. Nestes últimos, por sua vez, foi demonstrada correlação significativa da velocidade de eluição do isótopo no ...Observação: O resumo, na integra, podera ser visualizado no texto completo da tese digital Abstract: The causes of leg ulceration in the sickIe cell diseases are not known, although a role of vasooclusion is generally assumed. In this study we demonstrated differences of the clearence of Tc99m from the calf muscle of sickle cell anemia patients as compared with normals and patients with acquired anemias, which indicate the occurrence of abnormalities of peripheral blood flow in sickle cell anemia (SCA), 20 controls without anemia and vascular disease (N) and 20 patients with acquired anemia (AA); 20 patients with peripheral arterial disease (CI) were studied for method standardization. The clearence rates for the SCA and AA groups at rest were higher than for the controls. This difference, however, was significant only for SCA, and for this group there was a negative correlation between hemoglobin levels and clearence rate at rest. These results are probably the consequence of the decrease of the peripheral resistance in anemia. After exercise Tc99m clearence rate was significantly higler for controls N than for the other groups, which seems to reflect the lower increment of cardiac output of anemic patients. The peripheral vascular reserve (maximal clearence rate/clearence rate at rest) was significantly lower for patients Doutorado Doutor em Medicina

Loss of endoplasmic reticulum (ER) homeostasis leads to ER stress, thus prolonged activation can lead to apoptosis. Herein, artesunate (ART) induced ER stress in leukaemia cells, resulting in eIF2a phosphorylation, activation of transcription factor 4, subsequent CHOP upregulation and XBP1 splicing. Furthermore, in vitro cyclin/CDKs reduction induced G1-phase arrest. An in vivo xenograft model showed a consistent pattern of ART in reducing tumour burden, supporting roles in the UPR pathway, which we speculate could lead to apoptosis by NOXA activation. Moreover, ART were capable of increasing the survival of mice. Taken together, our data indicate that ART may represent an interesting weapon to fight leukaemia CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQ FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP Aberto

Agradecimentos: Te authors would like to thank Raquel S. Foglio for English review, and Márcio Alves da Cruz for his technical assistance with the animal’s care. Molecular structure of EGCG was produced using ChemDraw 19.0 sofware. Tis work was funded by the Conselho Nacional de Desenvolvimento Científco e Tecnológico (CNPq/MCT) [Grant Number 303405/2018-0]; the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) [Grant Numbers 11/51959-0 and 17/21801-2]; and the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES) [Finance Code 001] Abstract: (–)-Epigallocatechin-3-gallate (EGCG), the major active polyphenol extracted from green tea, has been shown to induce apoptosis and inhibit cell proliferation, cell invasion, angiogenesis and metastasis. Herein, we evaluated the in vivo efects of EGCG in acute myeloid leukaemia (AML) using an acute promyelocytic leukaemia (APL) experimental model (PML/RARa). Haematological analysis revealed that EGCG treatment reversed leucocytosis, anaemia and thrombocytopenia, and prolonged survival of PML/RARa mice. Notably, EGCG reduced leukaemia immature cells and promyelocytes in the bone marrow while increasing mature myeloid cells, possibly due to apoptosis increase and cell diferentiation. The reduction of promyelocytes and neutrophils/monocytes increase detected in the peripheral blood, in addition to the increased percentage of bone marrow cells with aggregated promyelocytic leukaemia (PML) bodies staining and decreased expression of PML-RAR oncoprotein corroborates our results. In addition, EGCG increased expression of neutrophil diferentiation markers such as CD11b, CD14, CD15 and CD66 in NB4 cells; and the combination of all-trans retinoic acid (ATRA) plus EGCG yield higher increase the expression of CD15 marker. These fndings could be explained by a decrease of peptidyl-prolyl isomerase NIMA-interacting 1 (PIN1) expression and reactive oxygen species (ROS) increase. EGCG also decreased expression of substrate oncoproteins for PIN1 (including cyclin D1, NF-?B p65, c-MYC, and AKT) and 67 kDa laminin receptor (67LR) in the bone marrow cells. Moreover, EGCG showed inhibition of ROS production in NB4 cells in the presence of N-acetyl-L-cysteine (NAC), as well as a partial blockage of neutrophil diferentiation and apoptosis, indicating that EGCG-activities involve/or are in response of oxidative stress. Furthermore, apoptosis of spleen cells was supported by increasing expression of BAD and BAX, parallel to BCL-2 and c-MYC decrease. The reduction of spleen weights of PML/RARa mice, as well as apoptosis induced by EGCG in NB4 cells in a dose-dependent manner confrms this assumption. Our results support further evaluation of EGCG in clinical trials for AML, since EGCG could represent a promising option for AML patient ineligible for current mainstay treatments CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO (CNPQ) FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO (FAPESP) COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPES Aberto

Agradecimentos: The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This work was supported by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq/MCT) [grant number PQ 301676/2013-5]; the Fundação de Amparo à Pesquisa do Estado de São Paulo [grant numbers 11/51959-0, 12/06675-7]; and the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Abstract: Green tea (GT) treatment was evaluated for its effect on the immune and antineoplastic response of elderly acute myeloid leukemia patients with myelodysplasia-related changes (AML-MRC) who are ineligible for aggressive chemotherapy and bone marrow transplants. The eligible patients enrolled in the study (n = 10) received oral doses of GT extract (1000 mg/day) alone or combined with low-dose cytarabine chemotherapy for at least 6months and/or until progression. Bone marrow (BM) and peripheral blood (PB) were evaluated monthly. Median survival was increased as compared to the control cohort, though not statistically different. Interestingly, improvements in the immunological profile of patients were found. After 30 days, an activated and cytotoxic phenotype was detected: GT increased total and naïve/effector CD8+ T cells, perforin+/granzyme B+ natural killer cells, monocytes, and classical monocytes with increased reactive oxygen species (ROS) production. A reduction in the immunosuppressive profile was also observed: GT reduced TGF-ß and IL-4 expression, and decreased regulatory T cell and CXCR4+ regulatory T cell frequencies. ROS levels and CXCR4 expression were reduced in bone marrow CD34+ cells, as well as nuclear factor erythroid 2–related factor 2 (NRF2) and hypoxia-inducible factor 1a (HIF-1a) expression in biopsies. Immune modulation induced by GT appears to occur, regardless of tumor burden, as soon as 30 days after intake and is maintained for up to 180 days, even in the presence of low-dose chemotherapy. This pilot study highlights that GT extracts are safe and could improve the immune system of elderly AML-MRC patients CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQ FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPES Aberto

Agradecimentos: This study was funded in part by the grant #2011/51959-0 and 2017/21801-2d, the São Paulo Research Foundation (FAPESP) and the Ministry of Science, Technology, Innovations and Communications (CNPq) grant # 301676/2013-5 Abstract: Inspiratory muscle training (IMT) has been shown to be an efficient method of improving exercise tolerance and inspiratory and expiratory muscle strength in several diseases. The effects of IMT on patients with sickle cell anemia (SCD) are relatively unknown. Our study aimed to evaluate the effects of IMT on adult SCD patients, regarding respiratory muscle strength (RMS) variables, lung function, exercise tolerance, blood lactation concentration, limitation imposed by dyspnea during daily activities and impact of fatigue on the quality of life. This was a randomized single-blind study, with an IMT design comprising true load (TG) and sham load (SG) groups. Initial assessment included spirometry, volumetric capnography (VCap) and measurement of RMS by maximal inspiratory and expiratory pressure (PImax and PEmax). The Medical Research Council dyspnea scale and modified fatigue impact scale were also applied and blood lactate concentration was measured before and after the 6-minute walk test. After this initial assessment, the patient used the IMT device at home daily, returning every 6 weeks for RMS reassessment. Both groups used the same device and were unaware of which group they were in. After a period totaling 18 weeks, patients underwent the final evaluation, as initially performed CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO (CNPQ) FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO (FAPESP) Aberto

Obesity is increasingly associated with the transformation of monoclonal gammopathy of undetermined significance (MGUS) into multiple myeloma (MM). Obesity, MGUS, and MM share common etiopathogenesis mechanisms including altered insulin axis and the action of inflammatory cytokines. Consistent with this interconnection, metformin could predominantly exert inhibition of these pathophysiological factors and thus be an attractive therapeutic option for MGUS. Despite the possible clinical significance, only a limited number of epidemiological studies have focused on obesity as a risk factor for MGUS and MM. This review describes multiple biological pathways modulated by metformin at the cellular level and their possible impacts on the biology of MGUS and its progression into MM.

Agradecimentos: This study was supported by São Paulo Research Foundation (FAPESP processes 17/21801-2, 2013/13022-2 and 17/19674-2), National Council for Scientific and Technological Development (CNPq) and Coordination for the Improvement of Higher Education Personnel (CAPES). The authors would like to thank Mariana Pissarra and Mariana Ozello Baratti for their valuable technical assistance. We also thank Raquel S Foglio for the English revision Abstract: Background: Microtubules, actin and Rho GTPase proteins are essential players in the megakaryocyte biology for platelet formation and function. Objectives: To investigate the role of ARHGAP21, a RhoGAP protein, in megakaryocytic differentiation and platelet function. Methods: Cytoskeletal proteins were investigated in HEL cells silenced for ARHGAP21 and submitted to megakaryocyte differentiation. The role of Arhgap21 in platelet function was accessed using haploinsufficient (Arhgap21+/-) mice. Arhgap21+/- platelet aggregation and p-selectin exposure were evaluated in response to thrombin. Vessel occlusion time and thrombus formation were detected after injury of the carotid artery. Platelet morphology was accessed by electronic microscopy. Results: ARHGAP21 was upregulated during megakaryocytic differentiation of the cell line and primary mouse cells. In the HEL cell model, ARHGAP21 was detected in the cytoplasmic protrusions, colocalized and associated with a-tubulin and was mostly detected in the protein cell fraction containing the polymerized tubulin. Silencing of ARHGAP21 decreased the expression of Glu-tubulin, suggesting microtubule instability, and enhanced cell spreading. Platelets from Arhgap21+/- mice presented enhanced thrombin-induced aggregation and p-selectin exposure associated with increased size of a-granules. Arhgap21+/- mice also showed increased CDC42 and RHOA activities, shorter tail-bleeding and accelerated thrombus formation. Conclusions: Our results indicate that ARHGAP21 may be a critical protein in the regulation of platelet production and function through the control of cytoskeletal rearrangement CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQ COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPES FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP Fechado

Despite being initially considered at higher risk for severe COVID-19, sickle cell disease (SCD) patients have mostly presented clinical severity similar to the general population. As their vulnerability to become infected remains uncertain, we assessed the seroreactivity for SARS-CoV-2 to estimate the prevalence of infection and possible phenotypic and socioeconomic determinants for their contagion. Serologic evaluation was performed on 135 patients with an overall prevalence of 11%; positivity was associated with older age and use of public transportation. We speculate that social distancing instructions recommended by our clinic may have contributed to lower levels of infection, but potential protection factors need further investigation FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP Aberto

Agradecimentos: This study was kindly funded by FAPESP # 2020/12622-0, #2019/25125-7, #2020/04133-9, Conselho Nacional de Desenvolvimento Científico e Tecnologico ' (CNPq), Programa de Apoio ao Desenvolvimento Científico – FCF-UNESP (PADC) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - Brasil (CAPES) - Finance Code 001 Abstract: Glioblastoma multiforme (GBM) is the most common primary brain cancer. GBM has aggressive development, and the pharmacological treatment remains a challenge due to GBM anatomical characteristics’ (the blood–brain barrier and tumor microenvironment) and the increasing resistance to marketed drugs, such as temozolomide (TMZ), the first-line drug for GBM treatment. Due to physical–chemical properties such as short half-life time and the increasing resistance shown by GBM cells, high doses and repeated administrations are necessary, leading to significant adverse events. This review will discuss the main molecular mechanisms of TMZ resistance and the use of functionalized nanocarriers as an efficient and safe strategy for TMZ delivery. GBM-targeting nanocarriers are an important tool for the treatment of GBM, demonstrating to improve the biopharmaceutical properties of TMZ and repurpose its use in anti-GBM therapy. Technical aspects of nanocarriers will be discussed, and biological models highlighting the advantages and effects of functionalization strategies in TMZ anti-GBM activity. Finally, conclusions regarding the main findings will be made in the context of new perspectives for the treatment of GBM using TMZ as a chemotherapy agent, improving the sensibility and biological anti-tumor effect of TMZ through functionalization strategies FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO (FAPESP) CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO (CNPQ) COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPES Fechado