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Background & Aim: During the COVID-19 pandemic, we performed HPC-A cryopreservation process validation using the CryoStor CS10 freeze media to replace the current 10% DMSO cryoprotectant (Control), which encountered severe backorder. Methods, Results & Conclusion(s): This process validation included phase I, phase II, and follow-up studies. Ten HPC-A collection cell product samples were cryopreserved in the phase I study using CS10 and Control (1:1) post-plasma depletion. Post-thaw viability tests using the 7-AAD method were performed on the cryopreserved samples for parallel comparison. In phase II, each of three patient HPC-A cell products was split evenly into CS10 and Control cryopreservation. The CS10 cryopreserved HPC-A cell products only were used for infusion. The recipients' engraftment outcomes of white blood cells (WBC), granulocytes (ANC), and platelets (Plts) were monitored. Post-thaw viability test was performed on the quality control samples from both groups. In the follow-up study, engraftment outcomes of WBC, ANC, and Plts were evaluated from ten recipients who received the CS10 cryopreserved HPC-A. In the phase I study, the post-thaw viability of the CS10 group was significantly higher than the Control group (p=0.002). All post-thaw viability results were above 60%, the current lab release criteria. In the phase II study, all cryopreserved cell products met cell product release criteria (> 60%). All engraftment results were within our center-established ranges except for the Pt b's platelet engraftment. Three recipients had not had any cell product infusion-related adverse events post infusion. Both CD34 and CD45 post-thaw viability results in the CS10 group were remarkably higher than the Control group, except for the patient c's CD34 viability. In the follow-up study, the total infused cell product volume ranged from 60 ml to 118 ml, and the WBC concentration in the cryopreserved cell products ranged from 134 to 440 (x10
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Introduction: Matched unrelated donors (MUD) for hematopoietic progenitor cell (HPC) transplantation are facilitated through the National Marrow Donor Program. Most peripheral blood collections (HPC-A) are obtained in a single day apheresis collection. Extensive planning is required to coordinate the mobilization, collection, and shipment of the product with the conditioning and infusing at the transplant center. Typically, these products are infused fresh, although the COVID pandemic has necessitated cryopreservation in many instances. It was perceived that the number of two-day MUD collections was increasing at our institution. This study was performed to determine if this was true and to evaluate potential causes. Method(s): The project was considered a laboratory quality improvement project;IRB approval was not required per institutional guideline. Data was collected retrospectively for 120 HPC(A) MUD from August 2017-November 2020 including donor's age, weight, and sex, along with recipient to donor weight ratio. Each factor was analyzed against CD34 yield per day of collection. Result(s): Of the 120 donors, 5.6% collected over 2 days in 2017(n=1), 3.7 % (n=1, 2018), 3.6 % (n=1, 2019) with highest observation 17% (n=8) in 2020 (Image). Donor age, donor weight, donor sex, and recipient to donor weight ratio were compared to absolute CD34 yield. There was not a correlation seen between CD34 yield and donor age nor weight. However, donor sex along with recipient/donor weight ratio each showed a correlation in the number of collections required. Of those requiring a second day of collection, 73% were female while 27% were male. Two-day collections could be predicted with 83% accuracy in female with >1.09 recipient/donor weight ratio and male with > 1.49 recipient/donor weight ratio.(Figure Presented) Conclusion(s): The observed trend of increased 2-day NMDP collections coincided with an increase in frequency of female donors. Not surprisingly, higher recipient/donor weight was associated with a higher likelihood of 2-day collections. The size and scope of this study do not allow us to determine a definitive cause. However, it was noted these findings coincided with new donor selection guidelines prioritizing HLA-DP match potentially leading to an increase in female donors being selected. Unexpected two-day collection can have significant effects on transplantation. Developing a predictive algorithm with 83% accuracy allows for patient and staff preparation to anticipate the likelihood for additional collections. Having the product collected and received in advance, prior to patient conditioning improves logistics and removes some variability from scheduling. Larger, multicenter studies are required to determine if increased numbers of two-day collection of MUD are occurring at other centers and to the potential causesCopyright © 2023 American Society for Transplantation and Cellular Therapy
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Background And Aims: S100A8 and S100A9 alarmins and their heterodimer calprotectin are diversely involved in myeloid neoplasm pathophysiology as well as infectious and inflammatory diseases. In the context of COVID-19, circulating calprotectin was identified as a powerful biomarker of disease severity. Calprotectin impact on CD34+ hematopoietic stem and progenitor cells remains poorly understood. Method(s): Calprotectin effects on healthy donor and chronic myeloid neoplasm-derived CD34-positive hematopoietic stem and progenitor cells were tested in liquid culture for up to 7 days. The pro-inflammatory cytokine IL-6 was used as a control. Cytokine effects alone or in combination were explored by the use of bulk and single cell RNA sequencing, Assay for Transposase-Accessible Chromatin with high-throughput sequencing, cytokine secretion analyses and semi-solid cultures. Result(s): CD34+ cells exposed to IL-6 generate monocytic cells that overproduce calprotectin. Calprotectin inhibits erythroid differentiation of healthy CD34+ cells, possibly through CD36 receptor. Chronic myeloid neoplasm CD34+ cells over-react to calprotectin, with large transcriptomic rewiring of erythro-megakarocytic and granulo-monocytic populations. Calprotectin-induced inhibition of erythroid progenitor proliferation correlates with increased synthesis of ribosomal subunits and p53 pathway activation, while the cytokine impact on granulo-monocytic cells indicates an autocrine or paracrine amplification loop. Conclusion(s): Calprotectin secreted by monocytes generated by CD34+ cells upon IL-6 stimulation may be a pathophysiological component of inflammatory anemia, a role that is amplified in the context of myeloid neoplasms in which calprotectin effects extend to the granulo-monocytic lineage.Copyright © 2023 Elsevier Ltd. All rights reserved.
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Introduction: Wolman disease is a rare genetic disorder with an autosomal recessive inheritance. A mutation in the LIPA gene causes lysosomal acid lipase (LAL) deficiency results in lipid storage and adrenal insufficiency. Death in early infancy is due to liver failure. Patients and methods: We describe the clinical course of a three-month-old infant diagnosed with Wolman disease. A rapid mutational analysis confirmed a LIPA gene defect. Results: He underwent matched unrelated donor peripheral blood stem cell hematopoietic stem cell transplantation (HSCT) at 3 months of age, with a treosulfan-based conditioning, which resulted in engraftment with donor-derived hematopoietic cells. He required supportive care for sinusoidal obstruction syndrome and mucositis. He was administered low dose prednisolone for grade I skin graft versus host disease, and a complete donor chimerism was documented on several occasions. At one year post HSCT, his growth and development were optimal, and there was no hepatosplenomegaly. He is maintained on glucocorticoid and mineralocorticoid supplements for primary hypoaldosteronism. Conclusion: The case emphasizes the timely diagnosis and the potential for successful treatment of Wolman disease by HSCT. © 2022 Pediatric Hematology Oncology Chapter of Indian Academy of Pediatrics
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Background: Allogeneic hematopoietic cell transplantation (HCT) with ex vivo T cell receptor (TCR) alphabeta+ T cell and CD19+ B cell depletion is an effective approach for children with primary immune deficiency disorders (PIDD) as it combines advantages of high CD34+ cell dose facilitating rapid engraftment with low risk of Graft Versus Host Disease (GVHD). The ideal pre-conditioning regimen that facilitates robust donor engraftment without increasing risk of transplant related mortality has not been well defined with this approach. Method(s): We report the outcomes of 4 pediatric subjects: Chronic Granulomatous Disease (CGD) (2), Wiskott Aldrich Syndrome (WAS) (1), and RAC2 deficient Severe Combined Immunodeficiency (1) who underwent haploidentical HCT with TCRalphabeta+ T cell/CD19+ depletion at Johns Hopkins All Children's Hospital/Moffitt Cancer Center from 2020-2022 (NCT04414046). Pre-conditioning regimen consisted of distal thymoglobulin (7.5 mg/kg), fludarabine (175 mg/m2), thiotepa (10 mg/kg) and pharmacokinetic guided busulfan targeting a cumulative area under curve (cAUC) (65-75 mgxhr/L). Rituximab (200 mg/m2) was administered on day +1. Result(s): The median age at HCT was 51 months (range 10-163 months). All patients received mobilized peripheral blood stem cells from HLA- haploidentical donors (paternal=1, maternal=1 sibling=2). Median busulfan cAUC for all patients was 69 mgxhr/L (range 65-76). Median CD34 and TCR alphabeta T cell dose was 9.13x106 cells/kg (range 7.0-18.9x106) and 0.7x105 cells/kg (range 0.09-1.0x105). Median times to neutrophil and platelet engraftment were 11 days (9-12) and 11 days (range 8-15), respectively. All 4 patients are alive with median follow-up of 19.5 months (range 7-24). One patient developed late VOD without organ dysfunction that resolved with defibrotide. At last follow up, peripheral T and myeloid chimerisms exceeded 90% in all 4 patients. Average time to CD4 recovery (> 200x106/L) was 142 days. Pre-existing inflammatory bowel disease in CGD (n=1) and WAS (n=1) patients resolved immediately following transplant. There was no graft failure, and none developed Grade III-IV acute or extensive chronic GVHD. Patient with WAS developed recurrent autoimmune cytopenias requiring corticosteroids, rituximab, sirolimus and daratumumab, and ultimately resolved. Viral reactivations included EBV (n= 1), adeno (n= 1), HHV6 (n= 2), BK (n=1), norovirus (n=1), and late HSV (n=1), all responded to antivirals without disease. All patients acquired SARS-Cov-2 after transplant and recovered without sequelae. Conclusion(s): TCR alphabeta+ and CD19+ depleted haploidentical transplantation using a reduced toxicity conditioning regimen with pharmacokinetic guided busulfan, fludarabine, thiotepa and thymoglobulin is well-tolerated in young children with PIDD that results in rapid, durable engraftment with low likelihood of GVHD and graft rejection.Copyright © 2023 American Society for Transplantation and Cellular Therapy
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Collection of peripheral blood stem cells (PBSCs) for autologous stem cell transplant (ASCT) requires mobilization from the bone marrow. There is variation in mobilization choice;during the COVID-19 pandemic BSBMT&CT guidelines recommended using granulocyte-colony stimulating factor (G-CSF) alone to minimize the use of chemotherapy. We report on the impact of mobilization regimen on stem cell collection, and whether IMiD-containing induction therapy impacts on mobilization and consequently transplant engraftment times for 83 patients undergoing ASCT at Leeds Teaching Hospitals. Cyclophosphamide plus G-CSF (cyclo-G) mobilization yielded more CD34+ cells (8.94 vs. 4.88 x106/kg, p = < 0.0001) over fewer days (1.6 vs. 2.4 days, p = 0.007), and required fewer doses of salvage Plerixafor than G-CSF only (13.6% vs. 35%, p = 0.0407). IMiD-containing induction impaired all of these factors. CD34+ doses > 8x106/kg were more frequent with Cyclo-G (62% vs. 11%, p = 0.0001), including for those receiving IMiD 1st line induction (50% vs. 13.3%, p = 0.0381). Note that 92.6% of those receiving IMiD-free inductions were mobilized with Cyclo-G. The novel agents used in modern induction regimens (e.g Daratumumab) have been shown to impair yields, increasing the importance of optimizing mobilization regimens in the first instance. Furthermore, as cellular therapies become established in the management of multiple myeloma emerging data highlights the potential benefits of stem cell top up in the management of the haematological toxicities of these therapies. Our findings support re-adoption of Cyclo-G as the gold standard for mobilization to optimize PBSC harvesting and ensure sufficient cells for subsequent ASCTs.Copyright © 2023 The Authors. eJHaem published by British Society for Haematology and John Wiley & Sons Ltd.
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Increasing number of severe COVID 19 patients develop pulmonary Fibrosis, but the management of this complication is still unclear due to a lack of clinical trials. Aim of this study was to characterize mesenchymal cells (MC) isolated from 10 broncho-alveolar lavage (BAL, at 2 months after discharge) from patients with COVID19 fibrosis (COVID19-f) and to compare them with those isolated from 8 patients with collagen tissue diseaseassociated interstitial fibrosis(CTD-ILD). BAL fluid (BALf) levels of TGFbeta, VEGF, TIMP2, RANTES, IL6, IL8, and PAI1 were assessed by ELISA. Primary MC foci were cultured and expanded in D-MEM +10% FBS, characterized by flow cytometry and osteogenic and adipogenic differentiation. Collagen 1 production (+/-TGF-beta) was tested by WB and mRNA expression. BALf cytokine and GF levels were comparable in the two groups. Efficiency of MC isolation from BAL was 100% in COVID-f compared to 65% in CTD-ILD. MC antigen surface expression of CD105, CD73, CD90 (>90%, respectively), CD45, CD34, CD19 and HLA-DR (<5%, respectively) was comparable. None of MC samples differentiated in adipocytes, while COVID19-f were positive for calcium deposition. COVID19-f MC showed at WB, higher Collagen 1 production with respect to CTD-ILD with TGF-beta stimulation. Our preliminary data suggest MC from COVID19-f share several features with CTD-ILD but might have a higher response to fibrogenic and differentiation signals.
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Current knowledge of histopathological changes in Covid-19 pneumonia is mainly based on autopsy findings. There are few data on dynamics of lung lesions in vivo after acute phase of disease. The aim of this study was to determine histopathologic changes during the long/post-Covid stage in patients who had suffered from moderate to severe Covid-19 pneumonia. Bronchoscopy with transbronchial lung biopsy was performed in patients with HRCT lesions involving >40% of lung parenchyma, at least 4 weeks after discharge. Additional criteria were restrictive pattern in lung function tests and signed informed consent. Histopathologic analyses were performed using H&E, MSB, MOVAT, TTF1, CD34 and CD68 staining. Research was approved by the Hospital Ethical Committee. Among 26 patients that met inclusion criteria, adequate biopsy samples were obtained from 24. The mean time from the onset of disease to biopsy was 13 weeks. We found 4 histopathologic patterns: diffuse alveolar damage-DAD with vascular abnormalities, nonspecific interstitial inflammation, organizing pneumonia and interstitial fibrosis in 11, 9, 2 and 2 patients, respectively. Vascular abnormalities included capillary thrombi, dilated venules and dissection of small pulmonary arteries. Given the duration of disease, DAD and vascular abnormalities were detected up to the 12 week from the onset of symptoms. All patients biopsied after 12th weeks had some degree of tissue inflammation without vascular changes. Our findings show rather slow recovery of lung tissue after Covid-19 pneumonia. Long lasting DAD with vascular abnormalities may explain prolonged dyspnea and exercise intolerance and should be taken into consideration when planning further rehabilitation.
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Objetivo: O presente trabalho tem como objetivo relatar um caso de evolucao de sindrome mielodisplasica para leucemia mieloide aguda, diagnosticada atraves de biopsia de lesao intraoral de sarcoma mieloide. Material e Metodos: Trata-se de um estudo descritivo, retrospectivo, do tipo relato de caso. Os dados foram coletados atraves dos registros em prontuarios eletronico e fisico da instituicao. Resultados: O caso apresentado envolveu um paciente do sexo masculino, de 51 anos, melanoderma, matriculado na instituicao devido ao diagnostico de sindrome mielodisplasica, tratado com 19 ciclos de decitabina. Devido a um quadro de odontalgia e abscessos de repeticao o paciente foi encaminhado ao setor de Odontologia onde, durante o exame clinico, foi constatada presenca de lesao eritematosa, amolecida, friavel, de superficie lisa, base sessil, de aproximadamente 3 x 1cm na regiao distopalatina do dente 26. No exame radiografico, foi observada imagem radiopaca associada a raiz mesial do dente 26, com margens definidas. Foi realizada biopsia excisional da lesao em regiao de palato com carater de urgencia e as hipoteses diagnosticas iniciais foram de granuloma piogenico e cisto radicular. O laudo histopatologico foi compativel com quadro de sarcoma mieloide comprometendo mucosa, com imuno-histoquimica positiva para CD34 e CD117 e negativa para CD163 e TdT. Diante do diagnostico previo de sindrome mielodisplasica, associado ao diagnostico de sarcoma mieloide oral, a equipe medica constatou a transformacao da doenca inicial em leucemia mieloide aguda. Foi definido novo protocolo terapeutico e realizado um ciclo de citorreducao com Citarabina (ara-C) subcutaneo e resgate com hidroxiureia + ara-c (HYDAC) + idarrubicina. Durante este ciclo, o paciente foi diagnosticado com Covid-19, evoluindo para obito, nao concluindo portanto, o protocolo terapeutico proposto. Discussao: O diagnostico de sarcoma mieloide oral permitiu que a equipe multiprofissional detectasse a transformacao da doenca primaria em leucemia mieloide aguda, resultando na reformulacao do tratamento e conduta do caso. O prognostico da leucemia mieloide aguda secundaria a sindrome mielodisplasica e pior quando comparado as primarias, bem como, apresenta baixas taxas de remissao apos quimioterapia intensiva e a sobrevida global mediana de 9-12 meses. O diagnostico dessas lesoes por parte do cirurgiao-dentista e considerado um desafio, visto que sao raras em cavidade oral e as apresentacoes clinicas costumam ser variadas e inespecificas. Conclusao: O presente relato de caso evidencia a importancia de uma avaliacao clinica minuciosa e do diagnostico preciso e completo de lesoes orais, que podem implicar diretamente na definicao da doenca de base, tratamento e prognostico do paciente. Alem disso, ratifica a importancia do cirurgiao-dentista na pratica clinica e na atuacao profissional de forma interdisciplinar e integrada com a equipe de saude. Copyright © 2022
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Os bancos de sangue de cordao umbilical e placentario (BSCUP), os laboratorios de processamento de medula ossea/ sangue periferico para transplante e os centros de tecnologia celular, passaram a receber a denominacao comum de Centros de Processamento Celular - CPC. O pais tem BSCUP 14 unidades publicas e 19 de natureza privada, totalizando 33 BSCUP. Uma analise retrospectiva dos relatorios da ANVISA identifica que em 2003 foram coletadas 26 unidades, com desqualificacao de 15,38% delas, o crescimento da coleta foi exponencial e dez anos depois, em 2013, foram coletadas 13.995 unidades (5,82% de desqualificacao). Em 2020, houve uma diminuicao expressiva de coletas, reflexo da pandemia de COVID-19: 4.918 unidades (desqualificacao de 12,69%). Este tipo de produtividade compromete a viabilidade financeira destes servicos, e encontrar formas de otimizar bolsas desqualificadas por volume ou quantidade de celulas para demais finalidades e uma vertente de gestao que deve ser estabelecida. O objetivo deste projeto foi a coleta de segmentos de cordao umbilical das unidades ja previamente validadas pelos BSCUP para extracao de celulas tronco, caracterizacao imunofenotipica e producao de meio condicionado isento de soro fetal bovino. A obtencao do meio condicionado (MC) da cultura de celulas tronco. Tem crescido cada vez mais o interesse pelo uso dos fatores de crescimento, citocinas e moleculas sinalizadoras livres no MC alem das vesiculas extracelulares, que se tornaram relevantes, tanto para diagnostico como para terapeutica, inclusive para aplicacoes oftalmologicas. Neste campo, identificamos a DOS que impacta profundamente a qualidade de vida das pessoas. Ha 15 anos o Laboratorio de Biologia Celular tem desenvolvido o soro autologo, para atendimento dos pacientes refratarios aos tratamentos convencionais e farmacologicos disponiveis, em especial aqueles pacientes submetidos ao Transplante de Medula Ossea e que desenvolveram DOS2ario a doenca de enxerto versus hospedeiro. No entanto, existem pacientes com impossibilidade de acesso venoso ou com sorologias reagentes para doencas infecciosas que sao impedidos de utilizar o produto. Diante disto, optou-se por produzir o MC de celulas tronco de cordao umbilical de parturientes jovens e sem comorbidades para obtencao do secretoma das celulas para avaliacao terapeutica na DOS. Um segmento do cordao umbilical foi retirado e processado seguido de plaqueamento e expansao para posterior identificacao de adesao ao plastico, caracterizacao imunofenotipica por citometria de fluxo utilizando marcadores como CD11b, CD13, CD14, CD34, CD31, CD36, CD45,CD73, CD90, CD 105, CD106 e HLA-DR. Todas as amostras tiveram adesao ao plastico com aspecto fibroblastoides e perfil imunofenpotipico corrobora com o determinado pela SITC. Para a obtencao de MC foram semeadas CTMcup ate 70% de confluencia e foram submetidas ao wash out, recebendo meio de cultura DMEM-F12 aditivado por 48 horas. Apos isto, foi coletado 60mL do secretoma das celulas para o experimento especificos in vitro. Copyright © 2022
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Introducao: A Leucemia Mieloide Aguda (LMA) e uma doenca agressiva, e em geral, de prognostico reservado. O tratamento com intuito curativo e realizado com altas doses de quimioterapia, seguido ou nao por transplante de medula ossea (TMO) alogenico. Porem, a recidiva ainda representa um desafio a ser superado, levando a busca de novas opcoes terapeuticas, como o inibidor de bcl-2, Venetoclax. Objetivo: Relatar o caso de um paciente jovem com LMA recidivada com resposta ao tratamento baseado em Venetoclax, seguido por TMO alogenico. Metodo: Levantamento de dados do prontuario e revisao da literatura. Relato do caso: VSV, sexo masculino, 21 anos, diagnosticado com LMA em 18/02/2020, mielograma com 60,4% blastos, com fenotipo CD45+ intermediario;CD34+;CD13+;CD33 parcial;CD64 parcial;HLA-DR parcial;MPO parcial e cariotipo 46, XY [20]. Sem possibilidade de avaliacao molecular da doenca. O paciente recebeu tratamento de inducao padrao (D3A7) atingindo remissao completa apos um ciclo e seguindo com consolidacao com 3 ciclos de ARA-C 3g/m2 ate maio de 2020 (optou-se por nao realizar o 4ciclo devido a pandemia por COVID19). Apresentava doenca residual minima (DRM) negativa em reavaliacao medular de agosto de 2020, mantendo-se ate junho de 2021, quando apresentou recidiva com 33,8% de blastos em imunofenotipagem de reavaliacao. Internado em julho de 2021 para QT de resgate com FLAG, evoluiu com choque septico e insuficiencia respiratoria, sendo necessario suporte em unidade de terapia intensiva. Apos recuperacao clinica, obteve novamente DRM negativa, sendo encaminhado para TMO alogenico aparentado (irma "full match"). Entretanto, enquanto aguardava o transplante, apresentou nova recidiva (IF com 5,4% de blastos), sendo optado por tratamento com Venetoclax + Azacitidina, com reducao de DRM para 1,5%. Recebeu o segundo ciclo de Venetoclax combinado com citarabina, devido a indisponibilidade da Azacitidina, seguido por DRM negativa. O paciente foi submetido ao TMO alogenico em 31/03/22, com avaliacao medular no D+60 com quimerismo de 100%, mantendo DRM negativa no D+120. Discussao: O uso do inibidor de BCL-2 (Venetoclax), em combinacao com agentes hipometilantes ou doses baixas de citarabina, foi aprovado para pacientes recem-diagnosticados com LMA inelegiveis para quimioterapia intensiva, revolucionando o tratamento da doenca. Publicacoes recentes vem demonstrando novos beneficios dessa associacao, tanto previamente ao TMO (em primeira linha ou com doenca recidivada), como na terapia de resgate para recidiva pos-TMO. Tambem tendo sido evidenciado sucesso terapeutico em faixas etarias menores (incluindo criancas). Esses novos estudos motivaram o uso de terapia baseada em venetoclax nesse paciente jovem, com doenca recidivada, conseguindo atingir e manter DRM negativa. Conclusao: O caso relatado demonstra eficacia do Venetoclax (em combinacao com hipometilante ou citarabina) em promover remissao profunda da doenca, tornando o paciente habil para o transplante e aumentando a probabilidade de sobrevida a longo prazo. Copyright © 2022
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Introducao: A remissao espontanea de leucemia mieloide aguda e um evento muito raro, e suas descricoes na literatura medica geralmente envolvem um evento infeccioso grave. Mais recentemente, a COVID-19 tambem foi descrita como um dos fatores associados a tal fenomeno, e a hipotese mais aceita para o mesmo e de que ha uma hiperativacao imune que tambem apresenta efeito antitumoral. Relato de caso: Paciente feminina, de 54 anos, foi admitida no Hospital Brigadeiro com historia de alteracao de nivel de consciencia e lesao em pododactilo direito que motivou a procura do pronto socorro. No hemograma da admissao, apresentava hemoglobina 8,2 g/dL VCM 87, plaquetometria 11.000/mm3, leucocitos 99560 com 88% de celulas de medio a grande tamanho, alta relacao nucleo citoplasma nucleolos evidentes e presenca frequente de bastonete de Auer. A imunofenotipagem de sangue periferico,22,4% destas celulas revelaram marcacao CD13, CD15++, CD33++/+++, CD34 par, CD38+++, CD64+, CD71+, CD117, CD123+, HLA-DR, MPO par, alem de 18,6% de monocitos maduros, com diagnostico de leucemia mieloide aguda. Foi colhido teste rapido para COVID-19 antes da internacao, com resultado positivo. Nos 3 dias que se seguiram, a paciente evoluiu com desconforto respiratorio, taquipneia, e a tomografia de torax revelou ainda broncopneumonia sobreposta a COVID-19, o que resultou em intubacao orotraqueal por insuficiencia respiratoria. Durante a intubacao, a paciente evoluiu com parada cardiorrespiratoria em atividade eletrica sem pulso, com retorno a circulacao espontanea apos 10 minutos. Dada a gravidade do quadro, foi optado por nao iniciar inducao quimioterapica. Ao longo da internacao em UTI, a paciente fez uso de piperacilina tazobactam com vancomicina nos primeiro 7 dias de internacao, teve Pseudomonas aeruginosa multissensivel isolada de aspirado traqueal, e recebeu meropenem e teicoplanina por novo evento de choque septico, por 7 dias. Sua internacao em UTI durou 41 dias, contudo, ao longo dos primeiros 30 dias de internacao houve clareamento de blastos em sangue periferico, e parametros como hemoglobina e plaquetometria tambem evoluiram com melhora. Ao final do 42degree dia, quando recebeu alta da UTI para a enfermaria, foi feita avaliacao medular, quel nao revelou blastos com as caracteristicas imunofenotipicas previamente descritas. A paciente se encontra atualmente internada e com remissao citomorfologica e sem doenca residual minima pela imunofenotipagem. Nao foi administrada qualquer terapia antineoplasica, somente hidroxiureia para fins de citorreducao, nos primeiros 7 dias do quadro. Discussao: O caso evidencia uma rara e ainda nao compreendida remissao de leucemia mieloide aguda apos infeccao por COVID-19 e complicacoes infecciosas bacterianas graves. Casos semelhantes ja foram descritos em sepse grave e COVID-19 isoladamente, mas a evolucao natural do quadro envolve a recaida da doenca apos alguns meses do quadro agudo. A terapia antineoplasica, por sua vez, e indispensavel para a remissao sustentada. A explicacao mais aceita para o fenomeno e de que um mecanismo imunologico ativado e producao excessiva de citocinas pro-inflamatorias, sobretudo na COVID-19, destruiria as celulas neoplasicas, que poderiam inclusive ser infectadas pelo virus. Conclusao: Apesar de o mecanismo exato da remissao ser desconhecido, o caso apresentado reforca o papel da imunidade do individuo no tratamento antineoplasico e justifica a imersao em mais estudos envolvendo imunoterapia para a leucemia mieloide aguda. Copyright © 2022
ABSTRACT
Background: Gemtuzumab ozogamicin (GO), an anti-CD33 immunoconjugate Antibody is currently approved in combination with 7 + 3 in low- and intermediate risk acute myeloid leukaemia (AML). These patients are candidate for consolidation with autologous stem cell transplantation (ASCT) particularly when MRD- is obtained. GO can improve the rate of MRD negativity. There are limited data on the effect of its addition on the mobilization of Hemopoietic Stem Cells (HSC). Aims: To assess the feasibility of mobilization of HSC after re-introduction into market of GO at 3mg/m2 in 2019. Methods: We retrospectively studied AML patients undergoing 3+7 + GO induction and Ara-C + Daunorubicine + GO, consolidation (doses are derived from label instructions and ALFA0701 study) and mobilization on day +20 using GCSF 10μg/kg. CD34+ were monitored, and patients were harvested when a threshold of 20 cells/μL was reached in peripheral blood. Results: In 2020 and 2021, also considering constrains caused by COVID-19 pandemics, we attempted mobilization in our 3 Italian centres of 14 patients with a diagnosis of CD33+ de novo-AML. The median age was 52 years (range 29-65 yrs.), 4 were males and 10 females;11 patients carried a mutation of NPM1 and all had a normal karyotype except one with t(10p12;11q14) (Table 1). All received 3+7+GO induction and achieved a CR. Therefore, we started consolidation (total ARA-C 8g/m2) + GO as inpatient. Ten patients (71%) reached the established threshold of 20 CD34+ /μL and were successfully harvested, while 4 patients (29%) failed mobilization. The median day of apheresis was D+26 from the start to chemotherapy (range 22- 39). The median number of circulating CD34+ cells on the day of collection was 35.9 cells/μL (range 20-2153 cells/μL). The median CD34+ harvested was 4.65 x 106/kg (range 1.8- 44.6 x 106/kg). In our cohort, 4 patients (28% of the entire cohort and 40% of the harvested patients) underwent ASCT, 3 achieved favourable engraftment, while in the last patient ASCT is ongoing. Several reasons prevented ASCT in the remaining 6 patients: 3 patients underwent allogeneic SCT (2 had positive MRD on harvested apheresis;1 was reclassified as high-risk ELN2017 due to RUNX1 mutation resulting from NGS panel), 2 refused ASCT and one suffered early relapse. Summary/Conclusion: In our patients, the addition of GO did not impair HSC mobilization and harvesting that was reached in about 71% of cases, similarly to the AML-10 trial of the EORTC and GIMEMA Leukemia Groups where 70% of patients were successfully harvested. Our data are particularly interesting because in the pivotal ALFA0701 study, only one patient underwent Autologous- SCT, but in the control arm. An important limit of our case-series is that only 4 patients were auto-transplanted, so we have scant data on engraftment. In particular, evaluating day to engraftment of platelets would be interesting, given the known increase of thrombocytopaenia in patients treated with GO. In conclusion, mobilization with GO is feasible and further studies are warranted to evaluate the effects of fractioned doses of GO on HSC mobilization and ASCT outcome;the ongoing trial GIMEMA AML1819 - EudraCT number 2019-003871-20 - will prospectively assess the effect of GO, but with lower doses of ARA-C (total ARA-C 6 g/m2). (Table Presented).
ABSTRACT
Background: Optimal consolidation for young R/R FL in the rituximab age remains uncertain and the benefit of ASCT is not clearly established. Aims: The FIL FLAZ12 trial (NCT01827605) is a prospective, multicenter, randomized, phase 3 trial, comparing RIT versus ASCT, as consolidation after chemoimmunotherapy, both followed by R maintenance in R/R FL. Methods: Pts aged 18-65 yrs, with R/R FL after 1 or 2 lines of chemoimmunotherapy, without significant comorbidities were enrolled. Patients received 3 courses of therapy chosen by the investigator among RCHOP, R-DHAP, R-FM, R-ICE, R-IEV or R-B. Pts achieving at least PR (according to Cheson et al. 2007) were randomized 1:1 to either RIT or ASCT before CD34+ collection. Conditioning for ASCT was BEAM or TEAM. RIT was given as previously described (Morschhauser et al., 2008). After consolidation, pts received R maintenance every 3 months for eight courses. Primary endpoint was PFS. Considering ASCT toxicity, it was hypothesized to be a superior choice, if capable of increasing 3-years PFS from 40% to 60% (two-side log-rank test with alpha of 5% and a power of 85%). Clinical secondary endpoints were ORR, CRR, OS, EFS and TTF. Results: Between Aug 2012, and Sep 2019, 164 pts were screened and 159 enrolled by 38 FIL Centers (enrolled population). Unfortunately, the study was prematurely closed due to low accrual. The data were analyzed on an ITT basis on May 2, 2021 with a median follow-up (mFU) from enrollment of 43 months and 75 PFS events. The two arms were clinically well balanced, with median age of 57 yrs (IQR 49-62), 55% male, 57% stage IV, 20% bulky disease. Tumor re-biopsy was performed in 79% pts. POD-24, retrospectively assessed was observed in 32% of pts. Two pts (1%) did not start treatment (non-confirmed histology and withdrawal). Sixteen (10%) pts discontinued before randomization (7 SD, 3 PD, 3 AE, 1 withdrawal, 2 poor compliance) and 141 (89%) were randomized to either RIT (71) or ASCT (70) (randomized population). Of these 19 (13%) (RIT 8, ASCT 11) did not receive the planned consolidation due to 7 PD, 4 AE, 1 medical decision, 2 poor mobilization, 2 withdrawals, 1 poor compliance, 2 protocol breaches, while 63 (89%) received RIT and 59 (84%) ASCT. After RIT, 61% of pts achieved CR and 23% PR, while after ASCT these were 70% and 9%. Estimated PFS at 3 yrs was 60% (95% CI: 46%-71%) in the RIT arm vs. 59% (95% CI:45%-70%) in the ASCT arm, p = 0.8613 (HR 0.96, 95%CI: 0.57,1.59). (Figure 1) 3yrs-OS was again superimposable in the two arms: 83% (95%CI: 69%-91%) in the RIT vs 85% (95% CI: 72%-91%) in the ASCT, p = 0.8310 (HR 1.10, 95%CI: 0.45,2.72). Grade ≥ 3 hematological toxicity was 46% in the RIT vs 94% in the ASCT arm (p < 0.001). For ASCT vs RIT grade ≥3 neutropenia occurred in 94% vs 41% of pts (p < 0.001). During follow-up, 4 pts died in remission: 1 AML (RIT), 2 SARS-COV2 infections (RIT) and 1 pneumonia (ASCT). Second cancers occurred in 3 pts after RIT and 7 after ASCT (p = 0.480). Multivariable analysis for PFS indicated POD-24, male sex, LDH and refractory disease as adverse parameters. Subgroup analysis for PFS including gender, age, LDH, POD-24 and extranodal disease show no subgroup favoring RIT nor ASCT. Image: Summary/Conclusion: Even if prematurely interrupted, our study demonstrated no meaningful difference in efficacy between ASCT and RIT, but ASCT was more toxic and more demanding for pts and health service. Both strategies induced a similar and favorable long-term outcome suggesting that consolidation programs milder than ASCT require further investigation in R/R FL.
ABSTRACT
Background: Second allogeneic hematopoietic cell transplant (sAHCT) might be indicated following a graft failure or disease relapse after the first one;although it might emerge with high rates of morbidities and mortality. Currently, there is a limited number of publications on this matter in the literature, here we aimed to share our sAHCT experience from a single center. Methods: Data from 51 patients who were eligible for sAHCT between 2001 and 2021 was evaluated retrospectively. All data was obtained from the Ankara University Faculty of Medicine, Department of Hematology and Bone Marrow Transplant Unit. Results: 51 patients were included in the present study. Median age at sAHCT was 34 (18- 65) and female/ male ratio 19/ 32 (37.3% / 62.7%). The same donor from the first transplant was eligible for sAHCT for most patients (n= 46, 90.2 %). sAHCT indication was graft failure for 11 patients (21.6 %) whereas 40 (78.4 %) patient went on sAHCT for disease relapse. Patients' diagnoses were as follows: acute myeloid leukemia (n= 26, 50.9 %), acute lymphoblastic leukemia (n=9, 17.6 %), myelodysplastic syndrome (n= 6, 11.8 %), aplastic anemia (n= 6, 11.8 %) and others (CMML, CML, biphenotypic leukemia). Median number of transplanted CD34+ hematopoietic cells was 5.77 x x106/ kg (1.11- 8.29). Stem cell source was either bone marrow (n= 5, 9.8%) or peripheral blood (n= 46, 90.2 %). Myeloablative conditioning regimens were used for most sAHCTs (n= 30, 58.8%). Median overall survival (OS) rates for graft failure and disease relapse groups were 12.8 and 18.7 months, respectively (p= 0.63). During early transplant phase, 20 patients (39.2 %) died due to bone marrow aplasia, transplant failure or other complications. 1 year OS of the entire cohort was 33.3 % whereas 2-y- OS was 21.6% (95% CI= 25-45). 2 patients (3.9 %) died due to COVID19 during transplant process. On univariate analysis, sex, time from the first transplant (<12 months/ ≥12 months), conditioning intensity, sAHCT indication did not statistically significantly influence OS. Multivariate analysis confirmed a lower ECOG score (<2) at sAHCT significantly increased OS (p= 0.001). Conclusions: Based on this single center study, sAHCT is an efficacious treatment modality especially for patients with lower ECOG scores. sAHCT may offer long term survival for both graft failure and disease relapse states.
ABSTRACT
The presence of coronavirus-associated myocarditis remains controversial despite elevations in cardiac troponin and natriuretic peptide in many patients. Aim. To assess the morphological changes in the myocardium of patients who died due to coronavirus disease 2019 (COVID-19) and compare them with the intravital level of cardiac biomarkers. Material and methods. A total of 420 hospital charts and 77 autopsies of those who died from COVID-19 were analyzed. In 15 of 77 cases (19%) with histologically suspected myocarditis, an immunohistochemical examination of the myocardium with antibodies to CD3, CD45, CD8, CD68, CD34, Ang1, VWF, VEGF, HLA-DR, MHC1, C1q, VP1 of enteroviruses was performed, and in 8 patients with immunohistochemically confirmed myocarditis (10%) — polymerase chain reaction for SARS-CoV-2. Results. Hemorrhage, intramural thrombosis, necrosis of non-coronary origin, myocardial infarction and lymphocytic myocarditis were detected in 43%, 10%, 17%, 19% and 10% of cases, respectively, without coronavirus N and E gene sequences in the myocardium. Dysplasia, hyperplasia and hypertrophy of the vascular endothelium, expression of Ang1, VWF, VEGF, MHC1, C1q, VP1 of enteroviruses were determined in 100, 100, 87, 100, 75 and 62% of cases of myocarditis, respectively. There were no significant correlations between inflammatory biomarkers and myo-carditis. Conclusion. The main morphological manifestation of COVID-19 in the myo-cardium is the so-called endotheliitis with dysplasia and endothelial activation, leading to hemorrhages, intramural thrombosis and necrosis. There is no con-vincing evidence of a direct involvement of coronavirus in myocarditis induction.
ABSTRACT
Background & Aim: Blood is one of the most vital resources in modern medicine. Blood transfusions have become an essential and often lifesaving procedure for accidents, during surgery, for patients with chronic disorders such as anemia, sickle cell disease, cancer, and myriad other circumstances. However, despite the rapidly growing world population, the availability of healthy blood donors is declining with aging populations. Furthermore, natural and man- made calamities often produce sudden and concentrated shocks in demand, which strains global supply chains. The COVID-19 pandemic has demonstrated this issue on a global scale by reducing the number of blood drives and donations, resulting in 39% of blood centers in the United States being left with only one- to two-day supplies, and a 50% drop of blood units collected in countries such as Zambia. Additionally, storage limitations of 42 days for donor blood limits stock availability during peak demand. Large-scale generation of universal red blood cells (RBCs) from O-ve human induced pluripotent stem cells (hiPSCs) offers the potential to alleviate blood shortages and provide a secure year-round supply. Mature iPSC-derived RBCs and reticulocytes could also find important applications in research in malaria and COVID-19 studies. (Figure Presented) Fig. 1 ( 700). Methods, Results & Conclusion: In this study, we have reprogrammed hiPSC from CD34+ O-ve cells and demonstrated the smallscale generation of high-density cultures of erythroblasts in a stirred perfusion bioreactor system. Twenty O-ve iPSC lines were derived, screened, and characterized for their ability to differentiate towards the erythroid lineage, showing high expression of mesoderm (KDR+, 64.9%), hematopoietic (CD34+/CD45+, 68.4%;CD34+/CD43+, 84.9%), and erythroid markers (CD235a+, 83,5%), and were able to undergo enucleation in vitro. Using the best clones, we were able to achieve erythroblast peak cell density of 34.7 million cells/mL with 92.2% viability in an Applikon perfusion bioreactor using an ultrasound system (Sonosep) to concentrate cells while removing waste media. This resulted in a cumulative-fold expansion of over 1,500 after 29 days of culture. Cells carried O2 effectively as demonstrated by hemoglobin dissociation curves. The perfusion culture platform paves the way for controlled high-density bioreactor culture for the generation of RBCs.
ABSTRACT
Background & Aim: Prior to the COVID-19 pandemic, allogeneic transplants were typically performed with fresh hematopoietic stem cell (HPSC) products. Unrelated donor (UD) cells are obtained through the National Marrow Donor Program (NMDP). The logistics for coordinating collection, transport, and delivery of fresh products with preconditioning of recipients is complicated under the best circumstances. The pandemic created uncertainty and disruptions in the UD HPSC process. In March 2020 the NMDP required cryopreservation of UD HPSC products, with rare exceptions, prior to patient conditioning. The impacts of cryopreservation on allogeneic HPSC engraftment are not well defined and conflicting outcomes based on transport time and cell concentration have been published. We aimed to determine if cryopreservation, transport time and pre-processing cell concentration negatively impacted patient engraftment. Methods, Results & Conclusion: Methods: Between July 2021 and January 2022, we analyzed UD HPSC products from 24 patients for CD34+ pre- and post-thaw cell recovery and viability based on transit time and pre-processing cell concentration. Transit time, defined as the interval from end of collection to start of processing, was divided into 3 cohorts: 1-20 h, 21-40 h, and >40 h. Pre-processing nucleated cell counts were divided into 2 cohorts: <200 x106 cells/mL and >200 x106 cells/mL. Neutrophil and platelet engraftment data were obtained from the patients’ medical record. Medians for 2 unpaired groups were compared by using Mann-Whitney U test. Three or more unpaired groups were compared using one-way ANOVA with Tukey’s multiple comparisons or Kruskal Wallis non-parametric test with Dunn’s test for post hoc analysis, as appropriate. For paired data a mixed model ANOVA with Geisser-Greenhouse correction was applied. Results: Information regarding patient diseases and product characteristics are shown in Table I. There were no statistically significant differences between the nucleated cell count in the product bag reported by the collection center and those measured at the time of processing. When these parameters were evaluated based on transit time and pre-processing cell concentration, no statistically significant differences were observed. Conclusion: Although our data set is small, the results suggest that transit time and cell concentration of the HPSC product bag does not negatively impact allograft quality and engraftment. (Table Presented)
ABSTRACT
Background & Aim: Background: Traditionally, ‘fresh’ Hematopoietic progenitors cell (HPC) infusions have been preferred over cryopreserved HPC in Allo-HCT because cryopreservation and thawing leads to cell loss, besides DMSO-related adverse reactions in patients. Emergence of COVID-19 pandemic has severely affected fresh HPC infusions and most professional bodies recommend cryopreservation of HPC products before initiating conditioning chemotherapy. Although some western studies suggest no significant impact of graft manipulation on patient outcome, there is no available data from the developing world.Aim: We compare neutrophil and platelet engraftment in patients undergoing Allo-HCT with fresh and cryopreserved HPC products. Methods, Results & Conclusion: Material and Method: Allo-HCT data from October 2018 to October 2021 were analyzed. Cryopreservation was performed by controlled-rate freezing using 10% DMSO, plasmalyte- A and human albumin ( 1:2:1) as cryoprotectant. Cryopreserved products were stored in vapour-phase of Liquid nitrogen tank. CD34+ enumeration and viablity( by 7-AAD) was done on Flow-cytometry on fresh and post-thaw HPC samples. Neutrophil engraftment was defined as absolute neutrophil count >0.5 ×109/L for 3 days. Platelet engraftment was defined as independence from platelet transfusion for at least 7 days with a platelet count >20 × 109/L. Statistical analysis using Wilcoxon Rank Sum test. Results: Ninety-six patients underwent allo-HCT (46 received fresh and 50 received cryopreserved HPC products) (Table 1). There was no significant difference in neutrophil engraftment with fresh and cryopreserved grafts (p>0.05) in different types of transplants( Matched related/unrelated and haploidentical). 22% (11/50) of cryopreserved graft infusions were associated with Grade-1 DMSO-related adverse reactions, which were managed with symptomatic treatment. Cryopreservation increased the cost of related allogeneic transplants by USD1100. No cryopreserved HPC product was culture positive on microbiological assessment. Conclusion: In our experience, the engraftment kinetics were similar with fresh and cryopreserved HPC products as CD34+cell dose administered was almost the same. Cryopreserved grafts had a median 7% CD34+cell loss, associated with mild DMSO-related adverse reactions and cost increment. Even though, graft cryopreservation is a feasible alternative during the pandemic, it is crucial to ensure graft quality and promptly manage DMSO-related adverse reactions.(Table Presented) Table 1 Comparison of Fresh and cryopreserved HPC products in Allo-HCT
ABSTRACT
Background & Aim: Hematopoietic progenitor cells (HPCs) are infused for hematopoietic reconstitution in the setting of malignancy and inherited or acquired hematological deficiencies. Given the global COVID-19 pandemic, the recommendation was made to cryopreserve all allogeneic HPCs to protect recipients by allowing for subclinical cases of infection to present prior to infusion. As such, consideration of HPC stability programs (SP) and their rigor has risen. The goal of a SP is to prove the rigor of a transplant program’s cryopreservation and storage standard operating procedures so that sufficient HPC viability and potency are maintained for engraftment. SPs are also required by accreditation agencies such as AABB and FACT. Many HPC SP have validated product expirations out to 10 years. Here we share our SP to 20 years with ongoing validation for 30-year expiration. Methods, Results & Conclusion: Program Design: Current testing frequency of our SP is within the first year, and then at three-year intervals (3, 6, 9, 12, 15, 19, 21). Our rolling SP includes 2 additional (Figure Presented) Fig. 1.Current vs proposed HPC product testing and cryopreseveration schema. samples tested at 0, 5, & 9 years, then at 3-year intervals (12, 15, 19, 21, 24, 27, 30). SP samples are collected from donors requiring additional days to reach their goal but are in excess at the conclusion of collection (e.g., Day 1 collection 4.5e6 CD34+ cells/Kg, goal 5e6 CD34+ cells/ Kg). Samples are collected on a quarterly basis with ten 1mL cryovials being drawn (Figure 1). CD3+ and CD34+ viabilities are tested after cryopreservation with an acceptable threshold set at ≥75% for both. Conclusion: We are validating our SP up to 20 years with intention to validate to 30 years. Thus far, our SP reveals product age has no to low correlation with engraftment, suggesting maintenance of potency over time in a cryopreservative of 10% DMSO, 10% plasma, and 30% PlasmaLyte-A with a final cell concentration of ≤3×108 NC/mL (Figure 2). Successful engraftment has been seen in all recipients. Transplant programs should modify testing frequency, acceptance criteria, and product expiration to meet individual need while working towards standardization in the field. Given the frequency of DLIs and 2nd/3rd transplants at the Mayo Clinic, a 30-year SP reflects the need of our transplant program.(Figure Presented)Fig. 2 . ANC and platelet engraftment dates for ≥10-year-old HPC products