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Introduction: Anesthetic strategies to improve postoperative pain management after Total Knee Arthroplasty (TKA) include multimodal pain management to minimize opiate requirements. Postoperatively, subcutaneous standard bupivacaine HCl is often used. Recent studies have suggested that high dose liposomal bupivacaine may result in improved outcomes. Objectives: This study aims to compare periarticular injection of 399mg (high dose) Liposomal Bupivacaine to Standard Bupivacaine in patients undergoing Total Knee Arthroplasty. Methods: We propose a phase II, single-institution, randomized, controlled, double-blinded, superiority trial with two parallel arms with a 1:1 allocation ratio. The two arms of the study will be standard bupivacaine and liposomal bupivacaine at 399mg dose. We will include patients with age >50 years, with primary knee osteoarthritis requiring TKA, ASA status 1 and 2, without opioid consumption in the last 3 months, and with signed informed consent. Exclusion criteria are history of substance abuse, uncontrolled psychiatric disorder, hepatic cirrhosis, renal failure, rheumatoid arthritis, immune arthritis, post-traumatic osteoarthritis, active malignancy or oncologic disease, pregnancy, synchronic surgical intervention, and morbid obesity with Body Mass Index (BMI) equal or greater than 40kg/m2. The primary outcome is time to first opioid rescue. Secondary outcomes include total opioid consumption, VAS scale, adverse events, GROC score, arc of motion, bupivacaine serum levels, length of stay, and mobilization. Discussion: Improving total knee arthroplasty postoperative pain without opioids decreases opioid-related side effects, reduces costs, and improves outcomes. By decreasing opiate use and complications related to them, Liposomal Bupivacaine may improve patient satisfaction as well as functional outcomes.

Introduction ATLL is a rare and aggressive neoplasm caused by the human T-lymphotropic virus type 1 (HTLV-1). It is estimated that at least 5-10 million people carry HTLV-1 and 2-5% out of them will develop ATLL. Our group demonstrated an increase of cells in G0/G1 phase of cell cycle and aneuploidy in CD4+ T-cells in HTLV-1 asymptomatic carriers. These findings may reflect an adverse intracellular environment, caused by genetic stress due to viral particles inserted into host DNA. Intracellular mechanisms aiming to control and prevent replication of cells carrying genetic aberrations in genes involved in cell cycle regulation, DNA repair and apoptosis could be activated to hold cell division while DNA damage is repaired. Based on this hypothesis, delay of cell cycle in G0/G1 may be a step in the process of oncogenesis. Herein, we did a pilot study in order to analyze the pattern of expression of a set of genes involved in cell cycle control and senescence in CD4+ T-lymphocytes of HTLV-1 infected individuals searching for additional genetic abnormalities in this setting. Methods Peripheral blood samples were tested obtained from five HTLV-1 asymptomatic carriers and four ATLL patients. T-CD4+ cells were isolated in magnetic column followed by RNA extraction. Subsequently, it was converted into cDNA for the assays of the qRT-PCR in microplates of 96 wells customized with 44 senescence related genes. A pool of five samples from healthy individuals (control group) was used as a calibrator. RNA transcription was measured using 7500 Fast real time PCR system (Applied Biosystems) and data were collected by the 7500 software v2.0.5 (Applied Biosystems). The expression levels of the target genes were calculated using the Livak and Schmittgen (2001) method. The mRNA expression was normalized using the β-glucuronidase (GUSB) gene (Applied Biosystems, cod. Hs99999908_m1) and those genes with expression ≥ 2x or ≤ 2x in comparison to control group were considered as differentially expressed and were chosen to be validated in a secondary cohort of thirty HTLV-1 infected individuals. Results In HTLV-1 asymptomatic carriers the median age was 55 years (37-62 years) and 20% (1/5) of them were males, in ATLL patients: 45 years (38-66 years) and 100% (4/4) were females, while the control group had median age of 43 years (22-62 years) and 60% (3/5) were males. Among the ATLL group, 50% (2/4) were classified as smoldering, 25% (1/4) were acute and 25% (1/4) were lymphomatous, according to Shimoyama classification. COL3A1, SPARC and TWIST1 genes were overexpressed and CDKN1A, ID1, IFNG and TERT were suppressed in HTLV-1 asymptomatic carriers when compared to healthy controls. HTLV-1 infected and ATLL patients showed TWIST1 and COL3A1 genes overexpressed and IFNG, CDKN1A and TERT repressed. The ALDH1, FN1, NOX4 and COL1A1 genes did not amplified. Conclusion These preliminaries results showed for the first time that HTLV-1 asymptomatic carriers present a set of genes differentially expressed in T-CD4+ cells, especially SPARC, TWIST1, COL3A1, CDNK1A and IFNG. These results will be confirmed in a validation cohort. We expected that this data shed some light on the comprehension of the cell microenvironment of the T-CD4+ lymphocyte in HTLV-1 asymptomatic carriers. Furthermore, we will be able to understand potential mechanisms associated with leukemogenesis in chronic infection by this virus, explaining eventual progression to ATLL.

Introduction: Adult T-cell leukemia/lymphoma (ATLL) is a rare and aggressive neoplasm caused by the human T-lymphotropic virus type 1 (HTLV-1). It is estimated, worldwide, that at least 5-10 million people carry HTLV-1 and 2-5% out of them will develop ATLL. Previously, our group demonstrated an increase of cells in G0/G1 phase of cell cycle and aneuploidy in CD4+ T-cells in HTLV-1 asymptomatic carriers [1]. These findings may reflect an adverse intracellular environment, caused by genetic stress due to viral particles inserted into host DNA. Intracellular mechanisms aiming to control and prevent replication of cells carrying genetic aberrations in genes involved in cell cycle regulation, DNA repair and apoptosis could be activated to hold cell division while DNA damage is repaired. Based on this hypothesis, delay of cell cycle in G0/G1 may be a step in the process of oncogenesis [1]. Herein, we did a pilot study in order to analyze the pattern of expression of a set of genes involved in cell cycle control and senescence in CD4+ T-lymphocytes of HTLV-1 infected individuals searching for additional genetic abnormalities in this setting. Methods: Peripheral blood samples were tested obtained from five HTLV-1 asymptomatic carriers and four ATLL patients for this pilot study. T-CD4+ cells were isolated in magnetic column followed by RNA extraction. Subsequently, it was converted into cDNA for the assays of the real-time quantitative polymerase chain reaction (qRT-PCR) in microplates of 96 wells previously customized with 44 senescence related genes pursued from ThermoFischer Scientific. A pool of five samples from healthy individuals (control group) was used as a calibrator. RNA transcription was measured using 7500 Fast real time PCR system (Applied Biosystems) and data were collected by the 7500 software v2.0.5 (Applied Biosystems). The expression levels of the target genes were calculated using the Livak and Schmittgen (2001) method [2]. The mRNA expression was normalized using the β-glucuronidase (GUSB) gene (Applied Biosystems, cod. Hs99999908_m1) and those genes with expression ≥ 2x or ≤ 2x in comparison to control group were considered as differentially expressed and were chosen to be validated in a secondary cohort of thirty HTLV-1 infected individuals. Results: In HTLV-1 asymptomatic carriers the median age was 55 years (37-62 years) and 20% (1/5) of them were males, in ATLL patients: 45 years (38-66 years) and 100% (4/4) were females, while the control group had median age of 43 years (22-62 years) and 60% (3/5) were males. Among the ATLL group, 50% (2/4) were classified as smoldering, 25% (1/4) were acute and 25% (1/4) were lymphomatous, according to Shimoyama classification [3]. COL3A1, SPARC and TWIST1 genes were overexpressed and CDKN1A, ID1, IFNG and TERT were suppressed in HTLV-1 asymptomatic carriers when compared to healthy controls. HTLV-1 infected and ATLL patients showed TWIST1 and COL3A1 genes overexpressed and IFNG, CDKN1A and TERT repressed (Figure 1 and 2). The ALDH1, FN1, NOX4 and COL1A1 genes did not amplified. Conclusion: These preliminaries results showed for the first time that HTLV-1 asymptomatic carriers present a set of genes differentially expressed in T-CD4+ cells, especially SPARC, TWIST1, COL3A1, CDNK1A and IFNG. These results will be confirmed in a validation cohort. We expected that this data shed some light on the comprehension of the cell microenvironment of the T-CD4+ lymphocyte in HTLV-1 asymptomatic carriers. Furthermore, we will be able to understand potential mechanisms associated with leukemogenesis in chronic infection by this virus, explaining eventual progression to ATLL. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Introduction: Isolation of circulating cell-free DNA (ccfDNA) and circulating tumor DNA (ctDNA) are useful methodologies for studies of neoplastic genetic material and can provide information about cancer heterogeneity, prognosis and minimal residual disease (MRD) [1,2]. Therefore, the recovery of ccfDNA in sufficient quantity and adequate purity is crucial to provide accurate and reproducible results. In this study, we investigate the efficiency of four different commercial kits to recover short fragments of DNA compatible with the ccfDNA. Methods: Ten mL of total peripheral blood from seven patients with Diffuse Large B-Cell Lymphoma (DLBCL) were collected in EDTA and were immediately processed. Plasma was obtained by centrifugation at 1.900 x g for 10 min at 4 ºC, followed by other centrifugation at 20.000 x g for 10 min at 4 ºC to remove cellular debris. Aliquots of 1.0 mL of plasma were stored at -80 ºC until ccfDNA extraction. The ccfDNA extraction was performed using the same samples by the following kits: QIAamp MinElute ccfDNA kit (CFDNA - Qiagen, Hilden, Germany), QIAamp DNA Mini Kit (QBLOOD - Qiagen, Hilden, Germany), Illustra Blood GenomicPrep Mini Spin Kit (GE - GE Healthcare Bio-Sciences Corp, UK) and ccfDNA isolation kit Quick-ccfDNA Serum & Plasma Kit (ZYMO - Zymo Research, Irvine, USA) [Table 1] according to the manufacturer's recommendations. The ccfDNA obtained was eluted in 30 µL of elution buffer for the subsequent experiments. The measurement of the ccfDNA obtained for each kit was performed in a Qubit 2.0 fluorometer using Qubit dsDNA HS assay kit (Thermo Fischer Scientific, Waltham, USA). The analysis of integrity, purity and the size of the fragments of DNA were performed in the Agilent 2100 Bioanalyzer (Agilent Technologies, Germany) equipment using the High Sensitivity DNA kit (Agilent Technologies, Germany). Results: The results obtained for each commercial kit in the Qubit-analysis were: CFDNA (0.27 ng/uL), QBLOOD (0.26 ng/uL), ZYMO (0.03 ng/uL) and GE (0.02 ng/uL) [Figure 1]. The Bionalyzer eletrophoresis showed that CFDNA kit was able to recover two different fragments sizes of ccfDNA one of 360-380 bp (0.114 ng/uL) and other of 160-180 bp (1.808 ng/uL). The DNA recovered with QBLOOD revealed sizes of 360-380 bp (0.031 ng/uL) and 160-180 bp (0.079 ng/uL). Both kits, GE and ZYMO were not able to isolate fragments of DNA smaller than 10.380 pb . Conclusion: We demonstrated that QIAamp MinElute ccfDNA kit (CFDNA) was capable to recovery high quantity of small fragments of DNA compatible with ccfDNA. This result supports the use of protocols based in magnetic beads in experiments working with ccfDNA. Additionally, we highlight the importance to verify the quality and not only the quantity of the DNA extracted to confirm the presence of ccfDNA. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Introduction:Adult T cell Leukemia/Lymphoma (ATLL) is a mature T-Cell-neoplasm related to human T-cell lymphotropic virus type 1 (HTLV-1) infection that shows variable clinical presentation and adverse prognosis with shorter overall survival (OS) when compared to other peripheral T-cell lymphomas (PTCL). Previous epidemiological studies estimated cumulative incidence in endemic regions around the world. In Brazil, mainly due to its vast dimension, data collection has limitations and is subject to bias. In April 2015 theBrazilianT-cell longitudinal project initiative was launched. One of the primary purposes was the collection of epidemiological and clinical data from the most frequent subtypes of newly diagnosed PTCL. Among them, 41 cases of ATLL were recorded. Objectives:The aim of this study is to describe clinical features, frequency and overall survival (OS) of 41 cases of ATLL registered in the ongoingBrazilianT-Cell Project. Methods:This is an ambispective observational study design collecting baseline characteristics, clinical features including date of diagnosis, clinical subtypes, B symptoms, performance status, Ann-Arbor staging, HTLV-1 status, number of sites, nodal and extra nodal presentation and types of skin lesions, peripheral blood counts and biochemical tests, front-line treatment and best response after first-line treatment. REDcap Platform has been used to collect and store data and for descriptive analysis the IBM-SPSS version 24 was applied. Kaplan-Meier method estimated the OS, whereas Log-Rank tests to compare its curves. OS period was calculated from diagnosis date until death or last seen date, and event was death by any cause. Results:Out of 281 cases of PTCL registered so far, 41 were ATLL cases. The median age was 50 years (34-88), 25 (64%) female; a higher incidence of lymphoma subtype was observed (46%), followed by acute (29%), chronic (17%) and smoldering (8%). Most of the patients (85%) had advanced-stage disease (III-IV, Ann-Arbor) and 56% had B symptoms (Table 1); 73% received chemotherapy with anthracycline-based regimens (46.5% CHOEP; 33.5% CHOP; 20% others) whereas 17% were managed with immunotherapy and antiviral therapy. The overall response rate was 30%; no response or progression 46%; stable disease 9% and 15% no data available yet (Table 2). Median follow-up was 13 months and 25 months for 41% of alive patients. Two year OS was 39% (95%CI: 23-55%) (Figure 1). Smoldering and Chronic subtypes showed better OS when compared with acute and lymphoma (100% smoldering, 86% chronic; 30% lymphoma type and 13% acuteP=0.04) (Figure 2). The multivariate Cox Regression analysis found male gender (HR 10.9 95%CI: 3.0-39.7, P Discussion:ATLL prognosis remains poor regardless of the type of treatment regimen and may be associated with the high incidence of lymphoma and acute subtypes, as well as advanced stage disease presentation. Despite the high number of cases seen in southeastern region of Brazil, it is important to emphasize there are still limited data from other regions. Albeit the sample size is small, these findings confirmed literature review data so far, with poor overall survival in a short time and gender and albumin as predictors of OS in the multivariate analysis. Conclusion:This study highlights the poor prognosis associated with ATLL. Moreover, it seems relevant to expand this study to all Brazilian regions. Hence, the need for early diagnosis and new treatment strategies, able to reduce mortality is warranted, that could possibly change the nature and spectrum of disease. Disclosures Federico: Spectrum:Consultancy, Membership on an entity's Board of Directors or advisory committees;Sandoz:Consultancy, Membership on an entity's Board of Directors or advisory committees;Cephalon/Teva:Research Funding;Mundipharma s.r.l.:Research Funding;Celgene:Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding;Millennium/Takeda:Research Funding;Roche:Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding;Takeda:Consultancy, Membership on an entity's Board of Directors or advisory committees.