RESUMO
BACKGROUND AIMS: The most widely accepted starting materials for chimeric antigen receptor T-cell manufacture are autologous CD3+ T cells obtained via the process of leukapheresis, also known as T-cell harvest. As this treatment modality gains momentum and apheresis units struggle to meet demand for harvest slots, strategies to streamline this critical step are warranted. METHODS: This retrospective review of 262 T-cell harvests, with a control cohort of healthy donors, analyzed the parameters impacting CD3+ T-cell yield in adults with B-cell malignancies. The overall aim was to design a novel predictive algorithm to guide the required processed blood volume (PBV) (L) on the apheresis machine to achieve a specific CD3+ target yield. RESULTS: Factors associated with CD3+ T-cell yield on multivariate analysis included peripheral blood CD3+ count (natural log, ×109/L), hematocrit (HCT) and PBV with coefficients of 0.86 (95% confidence interval [CI], 0.80-0.92, P < 0.001), 1.30 (95% CI, 0.51-2.08, P = 0.001) and 0.09 (95% CI, 0.07-0.11, P < 0.001), respectively. The authors' model, incorporating CD3+ cell count, HCT and PBV (L), with an adjusted R2 of 0.87 and root-mean-square error of 0.26 in the training dataset, was highly predictive of CD3+ cell yield in the testing dataset. An online application to estimate PBV using this algorithm can be accessed at https://cd3yield.shinyapps.io/cd3yield/. CONCLUSIONS: The authors propose a transferrable model that incorporates clinical and laboratory variables accessible pre-harvest for use across the field of T-cell therapy. Pending further validation, such a model may be used to generate an individual leukapheresis plan and streamline the process of cell harvest, a well-recognized bottleneck in the industry.
Assuntos
Receptores de Antígenos Quiméricos , Adulto , Humanos , Linfócitos T , Contagem de Células Sanguíneas , Transplante Autólogo , Leucaférese , AlgoritmosRESUMO
Poor graft function is a serious complication following allogeneic hematopoietic stem cell transplantation. Infusion of CD34+-selected stem cells without pre-conditioning has been used to correct poor graft function, but predictors of recovery are unclear. We report the outcome of 62 consecutive patients who had primary or secondary poor graft function who underwent a CD34+-selected stem cell infusion from the same donor without further conditioning. Forty-seven of 62 patients showed hematological improvement and became permanently transfusion and growth factor-independent. In multivariate analysis, parameters significantly associated with recovery were shared CMV seronegative status for recipient/donor, the absence of active infection and matched recipient/donor sex. Recovery was similar in patients with mixed and full donor chimerism. Five -year overall survival was 74.4% (95% CI 59-89) in patients demonstrating complete recovery, 16.7% (95% CI 3-46) in patients with partial recovery and 22.2% (CI 95% 5-47) in patients with no response. In patients with count recovery, those with poor graft function in 1-2 lineages had superior 5-year overall survival (93.8%, 95% CI 82-99) than those with tri-lineage failure (53%, 95% CI 34-88). New strategies including cytokine or agonist support, or second transplant need to be investigated in patients who do not recover.
Assuntos
Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Sobrevivência de Enxerto , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Doadores de Tecidos , Condicionamento Pré-Transplante , Transplante HomólogoRESUMO
Haematological engraftment was assessed in 804 autologous transplants. Neutrophil recovery occurred in over 99% within 14 d but platelet recovery was delayed beyond this time in 14·8%. Time to recovery was dependent on the progenitor cell dose infused. The minimum CD34+ cell threshold adopted in this study (2 × 106 /kg) was safe although recovery was faster with a dose >5 × 106 /kg. CD34+ cell doses of between 1 and 2 × 106 /kg were also acceptable if either the granulocyte-macrophage colony-forming cell dose exceeded 2 × 105 /kg or this dose was due to splitting a higher yield harvest. Prompt neutrophil recovery affords important quality assurance for laboratory processing.
Assuntos
Sobrevivência de Enxerto , Hematopoese , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas , Adolescente , Adulto , Idoso , Contagem de Células Sanguíneas , Feminino , Transplante de Células-Tronco Hematopoéticas/métodos , Transplante de Células-Tronco Hematopoéticas/normas , Humanos , Masculino , Pessoa de Meia-Idade , Garantia da Qualidade dos Cuidados de Saúde , Fatores de Tempo , Condicionamento Pré-Transplante , Transplante Autólogo , Resultado do Tratamento , Adulto JovemRESUMO
Standard quality assurance (QA) of cryopreserved peripheral blood stem cells (PBSC) uses post-thaw viable CD34(+) cell counts. In 2013, concerns arose at Great Ormond Street Hospital (GOSH) about 8 patients with delayed engraftment following myeloablative chemotherapy with cryopreserved cell rescue, despite adequate post-thaw viable cell counts in all cases. Root cause analysis was undertaken; investigations suggested the freeze process itself was a contributing factor to suboptimal engraftment. Experiments were undertaken in which a single PBSC product was divided into three and cryopreserved in parallel using a control-rate freezer (CRF) or passive freezing method (-80°C freezer) at GOSH, or the same passive freezing at another laboratory. Viable CD34(+) counts were equivalent and adequate in each. Granulocyte-monocyte colony-forming unit assays demonstrated colonies from the products cryopreserved using passive freezing (both laboratories), but no colonies from products cryopreserved using the CRF. The CRF was shown to be operating within manufacturer's specifications with freeze-profile within acceptable limits. This experience has important implications for quality assurance for all transplant programmes, particularly those using cryopreserved products. The failure of post-thaw viable CD34(+) counts, the most widely used routine QA test available, to ensure PBSC function is of great concern and should prompt reassessment of protocols and QA procedures.
Assuntos
Criopreservação , Células-Tronco de Sangue Periférico/citologia , Células-Tronco de Sangue Periférico/metabolismo , Antígenos CD34/metabolismo , Biomarcadores , Sobrevivência Celular , Ensaio de Unidades Formadoras de Colônias , Criopreservação/métodos , Sobrevivência de Enxerto , Humanos , Contagem de Leucócitos , Transplante de Células-Tronco de Sangue Periférico/métodos , Transplante de Células-Tronco de Sangue Periférico/normas , Garantia da Qualidade dos Cuidados de Saúde , Fatores de TempoRESUMO
High dose melphalan (HDM) followed by autologous stem cell transplantation (ASCT) remains the standard consolidation in transplant eligible multiple myeloma (MM) patients. The timing between HDM administration and hematopoietic stem cell return (HSCR) varies among institutions, with a 'rest period' of 48 hours (h) employed by some for patients with renal impairment (RI). We investigated the differences in hematopoietic recovery and HDM toxicity between MM patients with RI who had HSCR after 24 vs 48 h from HDM. Fifty MM patients with RI (48 h group; n = 31 and 24 h group; n = 19) were included. No statistically significant differences were noted in surrogates for hematopoietic recovery and HDM toxicity between both groups. Only one death occurred in the 24 h group. No patients required renal replacement therapy. Therefore, a 24 h period between HDM and AHSC infusion appears safe for MM patients with RI.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Mieloma Múltiplo , Humanos , Melfalan , Mieloma Múltiplo/complicações , Mieloma Múltiplo/diagnóstico , Mieloma Múltiplo/tratamento farmacológico , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Transplante AutólogoRESUMO
In vitro exposure of haematopoietic stem and progenitor cells (HSPC) to cytokines in expansion or gene therapy protocols reduces homing and engraftment in vivo. We have previously reported that this is related in part to altered tissue specificity of short-term homing, leading to loss of cells in non-haematopoietic tissues. Here we demonstrate that defective engraftment persists when cultured HSPC are transplanted by intrabone injection. Changes in engraftment function occur within 24 h of cytokine exposure, and are evident when engraftment is analysed solely in the injected bone. A novel ex vivo model of the bone marrow was developed, in which the attachment of infused HSPC in rodent long bones is reduced following culture with cytokines. Finally, cultured HSPC demonstrated reduced adhesion to N-cadherin, osteopontin and vascular cell-adhesion molecule-1, ligands present in bone marrow niches. These changes in adhesive function occur rapidly, and are not related to downregulation of the relevant receptors. Our findings suggest that cytokine exposure of adult human HSPC results in altered adhesion within bone marrow niches, further leading to reduced engraftment potential in vivo.
Assuntos
Medula Óssea/patologia , Adesão Celular/fisiologia , Fatores Estimuladores de Colônias/farmacologia , Sobrevivência de Enxerto/fisiologia , Mobilização de Células-Tronco Hematopoéticas , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/patologia , Nicho de Células-Tronco/fisiologia , Adulto , Animais , Antígenos CD34/análise , Células Cultivadas/efeitos dos fármacos , Células Cultivadas/transplante , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/farmacologia , Organismos Livres de Patógenos EspecíficosRESUMO
The role of donor lymphocyte infusion (DLI) in the management of lymphoid malignancies after allogeneic stem cell transplantation (SCT) has not been clearly characterized. There is emerging evidence pointing to the effectiveness of this approach, particularly in patients with low-grade disease, although to date this has been reported only in small numbers of patients, and thus the utility of this treatment remains uncertain. A total of 28 patients with low-grade lymphoid malignancies previously treated with allogeneic SCT received a total of 68 infusions of donor lymphocytes. The diagnoses were indolent non-Hodgkin lymphoma (NHL; n = 23) and transformed NHL (n = 5), and the indications for DLI were progressive disease with or without mixed chimerism (MC) (n = 17) and persistent MC alone (n = 11). Escalating doses of cells were administered in the absence of graft-versus-host disease (GVHD) or continued disease progression, until stable full donor chimerism or disease response were achieved. The cumulative response rates after DLI to treat progressive disease and persistent MC were 76.5% and 91.6%, respectively. The major toxicity resulting from the use of donor lymphocytes was GVHD. The cumulative incidence of acute grade II-IV disease was 15%, and that of extensive chronic disease was 31%; there were no deaths resulting from GVHD. Seven patients had graft-versus-lymphoma responses without significant GVHD. These data support the existence of a clinically significant graft-versus-tumor effect in indolent NHL and suggest that this is an effective treatment for progressive disease after allogeneic SCT.
Assuntos
Efeito Enxerto vs Tumor , Transplante de Células-Tronco Hematopoéticas/métodos , Transfusão de Linfócitos/métodos , Linfoma não Hodgkin/terapia , Terapia de Salvação/métodos , Quimeras de Transplante , Adulto , Intervalo Livre de Doença , Feminino , Doença Enxerto-Hospedeiro/imunologia , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/prevenção & controle , Estudos Retrospectivos , Transplante Homólogo/imunologia , Transplante Homólogo/métodosRESUMO
The nature of hemopoietic progenitors subject to leukemic transformation in acute myeloid leukemia (AML) has not been clearly defined. To address this issue, we have used DNase I hypersensitivity assays to study the chromatin structure surrounding the T-lineage-affiliated CD2 gene in the acute promyelocytic subtype of AML (APL). Upstream and downstream flanking regions of CD2 were found to be hypersensitive to DNase I in primary APL blasts, with an identical pattern of hypersensitive sites to those detected in cells of T-lineage. All of the sites were confirmed to be inaccessible to DNase I in B-lineage leukemia cells. The demonstration of T-cell-associated chromatin features in primary APL blasts has implications for the origin of APL that may arise in more primitive progenitors than previously considered to be the case.
Assuntos
Antígenos CD2/genética , Cromatina/fisiologia , Leucemia Promielocítica Aguda/genética , Linfócitos T/fisiologia , Linhagem da Célula , Cromatina/química , Cromatina/genética , Desoxirribonuclease I/metabolismo , Humanos , Imunofenotipagem , Células Jurkat , Leucemia Promielocítica Aguda/patologia , Reação em Cadeia da Polimerase/métodos , Linfócitos T/citologiaAssuntos
COVID-19 , Transplante de Células-Tronco Hematopoéticas , Mieloma Múltiplo , Células-Tronco de Sangue Periférico , Antígenos CD34 , Fator Estimulador de Colônias de Granulócitos , Mobilização de Células-Tronco Hematopoéticas , Humanos , Mieloma Múltiplo/terapia , SARS-CoV-2 , Transplante AutólogoRESUMO
Mobilised peripheral blood is now the main source of stem cells collected from normal donors. We report our experience of mobilising and collecting 400 normal healthy donors using standardised procedures and techniques. Target recipient doses were reached with one aphaeresis in 63% of donors and with two aphereses in 81% of donors. Approximately 2% of donors yielded such low progenitor values that they were termed 'poor mobilisers'. There were minor effects of donor age, weight and sex and where possible, larger male donors under the age of 55 years should be selected. Two forms of granulocyte colony-stimulating factor (G-CSF) were used at the same dose and no significant difference was seen in the yield of CD34+ cells collected/l of blood processed. However, a greater number of granulocyte-macrophage colony-forming cells were harvested using lenograstim (glycosylated G-CSF) compared with filgrastim (non-glycosylated G-CSF; P = 0.002). CD34+ cell yields were also measured halfway through the aphaeresis procedure. This was found to be highly predictive of final yield and facilitated distribution of the stem cell product to other centres. The observation that CD34+ yields did not decline in the second half compared with the first half of aphaeresis suggests that the circulating cell numbers are not static.
Assuntos
Remoção de Componentes Sanguíneos/métodos , Seleção do Doador/normas , Fator Estimulador de Colônias de Granulócitos/farmacologia , Mobilização de Células-Tronco Hematopoéticas/métodos , Células-Tronco Hematopoéticas , Doadores Vivos , Adolescente , Adulto , Fatores Etários , Idoso , Remoção de Componentes Sanguíneos/instrumentação , Peso Corporal , Contagem de Células , Seleção do Doador/métodos , Feminino , Citometria de Fluxo , Humanos , Lenograstim , Contagem de Leucócitos , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Transplante de Células-Tronco de Sangue Periférico , Proteínas Recombinantes/farmacologia , Fatores SexuaisRESUMO
The two clinical scale devices currently available for CD34+ cell selection from peripheral blood stem cells (PBSC) apheresis products, the CliniMACS and the Isolex 300i, were compared directly by pooling and splitting two PBSC harvests collected on sequential days from 10 patients and processing half of each pooled harvest on each device. The CliniMACS product had significantly higher median CD34+ purity (90%vs 78%; P = 0.004) and lower median T-cell content (0.06%vs 0.44%; P = 0.003) compared with the Isolex 300i product. The median CD34+ yields were similar (64% and 60% respectively). However, when the functional capacities of the products were compared, the median recovery of colony-forming units was significantly greater from the Isolex 300i product (48%vs 38%; P = 0.035), as was expansion of cells in either erythroid or granulocytic lineage-specific liquid culture (2.1-fold more erythroid and 1.5-fold more granulocytic lineage progenitors on d 9 (P = 0.03 and 0.03 respectively). This was due to a higher proportion of apoptotic cells in the CliniMACS product (28%vs 18%; P = 0.007, annexin V binding). Hence, although the CliniMACS device yielded a higher purity product with fewer T cells, the Isolex 300i product contained fewer apoptotic cells and consequently had greater functional capacity in culture.
Assuntos
Antígenos CD34 , Remoção de Componentes Sanguíneos/instrumentação , Linfócitos/imunologia , Células-Tronco , Linfócitos B , Movimento Celular , Células Clonais , Humanos , Contagem de LinfócitosRESUMO
The reduced engraftment potential of hematopoietic stem/progenitor cells (HSPCs) after exposure to cytokines may be related to the impaired homing ability of actively cycling cells. We tested this hypothesis by quantifying the short-term homing of human adult CD34+ cells in nonobese diabetic/severe combined immunodeficient (NOD/SCID) animals. We show that the loss of engraftment ability of cytokine-activated CD34+ cells is associated with a reduction in homing of colony-forming cells (CFCs) to bone marrow (BM) at 24 hours after transplantation (from median 2.8% [range, 1.9%-6.1%] to 0.3% [0.0%-0.7%]; n = 3; P < .01), coincident with an increase in CFC accumulation in the lungs (P < .01). Impaired BM homing of cytokine-activated cells was not restored by using sorted cells in G0G1 or by inducing cell cycle arrest at the G1/S border. Blocking Fas ligation in vivo did not increase the BM homing of cultured cells. Finally, we tested cytokine combinations or culture conditions previously reported to restore the engraftment of cultured cells but did not find that any of these was able to reverse the changes in homing behavior of cytokine-exposed cells. We suggest that these changes in homing and, as a consequence, engraftment result from the increased migratory capacity of infused activated cells, leading to the loss of selectivity of the homing process.
Assuntos
Células da Medula Óssea/citologia , Movimento Celular/imunologia , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Imunodeficiência Combinada Severa/patologia , Animais , Antígenos CD34/análise , Células da Medula Óssea/química , Ciclo Celular/imunologia , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Quimiocina CXCL12 , Quimiocinas CXC/farmacologia , Meios de Cultura/farmacologia , Citocinas/fisiologia , Proteína Ligante Fas , Citometria de Fluxo , Células-Tronco Hematopoéticas/química , Humanos , Glicoproteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCIDRESUMO
Myeloma remains incurable with a median survival of 4 years, but outcome can be improved by the use of high-dose therapy. We used the etoposide, methylprednisolone, cytarabine and cisplatin (ESHAP) regimen as second-line therapy in 42 newly diagnosed myeloma patients who had failed vincristine, adriamycin and dexamethasone (VAD)- type therapy (n = 36), responded to first-line treatment but persisted in having significant residual marrow plasmacytosis (n = 5) or failed prior stem cell harvesting (n = 1), with the dual aim of improving disease response and mobilizing peripheral blood stem cells. Fourteen of 21 (67%) patients with no change or progressive disease after VAD responded to ESHAP; seven of 12 (58%) patients with minor response converted to partial response. Marrow plasmacytosis fell from a median of 52% at diagnosis to 23.5% after primary therapy and to15% after ESHAP. ESHAP chemotherapy was well-tolerated. There were 11 admissions due to febrile neutropenia (n = 7), nausea and vomiting (n = 2), pneumonia (n = 1) and perforated bowel (n = 1). Renal function deteriorated in 13 of 42 patients after ESHAP, but none required renal support. ESHAP mobilization was performed in 32 patients of whom 87% achieved a CD34(+) yield >2 x 10(6)/kg. In all, 38 patients proceeded to high-dose therapy. The overall survival for all patients was 62% at 4 years following ESHAP. We conclude that ESHAP has acceptable toxicity and efficient stem cell mobilizing capability, effectively cytoreduced this chemoresistant group of patients, and did not appear to adversely affect transplant outcome.