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1.
Transplant Cell Ther ; 29(6): 396.e1-396.e5, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-36828177

RESUMO

BACKGROUND: Extracorporeal photopheresis (ECP) is frequently utilized in the treatment of steroid-refractory acute and chronic graft-versus-host disease (GVHD). Although the mechanism of action is not fully understood, it has been postulated that its therapeutic effect is immunologic tolerance linked to the associated apoptosis of the treated cells. Despite significant advances in allogeneic hematopoietic stem cell transplantation (HSCT), prophylaxis and treatment of GVHD remain a challenge and major limitation associated with this therapy. Use of ECP is a valuable strategy; however, it is time, cost, resource intensive, and not readily accessible. OBJECTIVE: In an effort to expand access to this therapy, we are investigating the use of cryopreserved ECP-treated cells. This will provide the ability to administer a significant proportion of the treatment at a facility closer to the patient's residence, thereby decreasing the number of visits to the primary treatment center with the goal of improving and expanding access to this therapy. Here we report the effects of cryopreservation on ECP-treated leukocytes. STUDY DESIGN: Mononuclear cells were pheresed from human patients, ECP-treated, and collected for viability and apoptotic analysis. Cells were then cryopreserved at -80°C or -150°C for 1 week, 1 month, and 3 months. Following thaw, repeat viability and apoptosis studies were performed on the leukocytes. RESULTS: WBC viability for freshly ECP-treated leukocytes was 84.5% ± 3.5 at 1 week, 87.3% ± 5.2 at 1 month, and 79.1% ± 1.1 at 3 months post thaw. Similar results were seen for cells frozen in cryovials. Leukocytes frozen the day after ECP treatment had 1 week and 1 month WBC viabilities of 84.0 ± 4.1 and 83.1 ± 2.1, respectively. Apoptotic potential was well preserved at 3 months, with cryopreserved ECP-treated lymphocytes being 19.2%, 44.5%, 75.5%, and 94.0% apoptotic after thaw on days 0, 1, 2, and 3 in culture, respectively. CONCLUSIONS: ECP-treated leukocytes cryopreserved at -80°C or -150°C for 3 months remain viable and as capable of apoptosis as freshly treated cells. Cryopreservation of an ECP-product warrants further in vivo investigation as a strategy to facilitate access to this needed therapy.


Assuntos
Doença Enxerto-Hospedeiro , Fotoferese , Humanos , Criopreservação , Doença Enxerto-Hospedeiro/prevenção & controle , Leucócitos , Linfócitos
2.
J Clin Apher ; 35(4): 342-350, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32640498

RESUMO

BACKGROUND: A new protocol has been developed on the Amicus Separator that enables the device to perform online extracorporeal photopheresis (ECP) procedures when used in conjunction with the Phelix photoactivation device and associated disposable kit. The objective of this study was to evaluate the safety and performance of the Amicus ECP System in adult subjects with steroid-refractory or dependent chronic graft vs host disease (cGVHD). STUDY DESIGN AND METHODS: Eight subjects with mild to severe cGVHD underwent 31 procedures. Subject safety evaluations were performed pre and post procedure and adverse events (AEs) were recorded during treatment and 24 hours after the last procedure. In vitro evaluations of the treated cells included hematology counts and lymphocyte apoptosis, viability and proliferation as measures for ECP procedure validation. RESULTS: For n = 23 evaluable procedures, median (range) procedure time was 88 (78-110) minutes, during which 2.9 (0.6-4.7) × 109 TNCs (approximately 90% MNCs) were treated and reinfused to the subjects. All subject safety evaluations (vitals, cell counts, plasma hemoglobin and bacterial and endotoxin testing) were within expected ranges. All device or procedure related AEs were mild in nature. After 24 hours in culture, 86 (52-98)% of treated lymphocytes were apoptotic compared to 27 (15-51)% in controls. Inhibition of lymphocyte proliferation was >91% in all procedures. CONCLUSION: ECP procedures were safely completed in adult subjects with SR-cGVHD treated using the new online Amicus ECP system.


Assuntos
Doença Enxerto-Hospedeiro/terapia , Fotoferese/métodos , Esteroides/uso terapêutico , Adulto , Idoso , Apoptose , Proliferação de Células , Sobrevivência Celular , Feminino , Taxa de Filtração Glomerular , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Técnicas In Vitro , Internet , Linfócitos/citologia , Masculino , Pessoa de Meia-Idade , Segurança do Paciente , Fotoquímica , Projetos Piloto , Risco , Resultado do Tratamento
3.
J Clin Apher ; 30(3): 154-61, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25213780

RESUMO

BACKGROUND: The ability to cryopreserve a portion of the cells treated during extracorporeal photopheresis (ECP) would improve therapy logistics, particularly for pediatric patients, by allowing multiple therapeutic doses to be collected from a single apheresis session. However, the effect of cryopreservation on ECP-treated cells is unknown (e.g., ECP-induced lymphocyte apoptosis and inhibition of proliferation). STUDY DESIGN AND METHODS: Mononuclear cell (MNC) apheresis products collected from healthy subjects were ECP-treated using offline methods. Fresh samples of ECP-treated and control cells were placed immediately in culture. The remainder of the cells were frozen in cryovials (n = 8) or cryobags (n = 8) at -80°C. After 1 week of -80°C storage, ECP-treated and control cells were thawed rapidly and samples were placed in culture. Lymphocyte apoptosis was assessed by phosphatidylserine exposure using Annexin V/7-AAD labeling. Lymphocyte proliferation after 3 days culture was measured using the carboxyfluorescein succinimidyl ester labeling technique. RESULTS: On Day 0, apoptosis levels were <5% in fresh ECP-treated and control cells and approximately 20% on thawing of cryopreserved ECP-treated and control cells. Apoptosis levels were comparable between the two cryopreserved groups immediately on thawing, indicating that ECP-treated cells were no more sensitive to the cryopreservation process than control cells. During 72-h culture, apoptosis levels increased to >80% in fresh and cryopreserved ECP-treated cells but remained near constant in both control groups. Inhibition of lymphocyte proliferation was >95% in all ECP-treated cells with no significant difference between fresh and cryopreserved cells (P = 0.12). CONCLUSION: Cryopreservation did not impair the apoptotic response or anti-proliferative effect of ECP-treated lymphocytes, thereby demonstrating early feasibility of this approach.


Assuntos
Apoptose , Remoção de Componentes Sanguíneos/métodos , Criopreservação , Linfócitos/citologia , Fotoferese/métodos , Anexina A5/química , Proliferação de Células , Separação Celular , Citometria de Fluxo , Fluoresceínas/química , Humanos , Leucócitos Mononucleares/citologia , Fosfatidilserinas/química , Succinimidas/química
4.
Transfusion ; 51(5): 1079-85, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21077911

RESUMO

BACKGROUND: Our objective was to determine the growth kinetics of bacteria in leukoreduced apheresis platelets (LR-AP) in a platelet (PLT) additive solution (PAS; InterSol, Fenwal, Inc.) compared to LR-AP stored in plasma. STUDY DESIGN AND METHODS: Hyperconcentrated, double-dose LR-AP were collected from healthy donors with a separator (AMICUS, Fenwal, Inc.). LR-AP were evenly divided, InterSol was added to half (65% InterSol:35% plasma [PAS]), and PLTs in autologous plasma were used for a paired control (PL). Bacteria were inoculated into each LR-AP PAS/PL pair (0.5-1.6 colony-forming units [CFUs]/mL), and bacterial growth was followed for up to 7 days. Time to the end of the lag phase, doubling times, maximum concentration (conc-max), and time to maximum concentration (time-max) were estimated. RESULTS: Streptococcus viridans did not grow to detectable levels in either PAS or PL units. The other bacteria had no significant overall difference in the conc-max (p = 0.47) or time-max (p = 0.7) between PL and PAS LR-AP; PL had a 0.14 hours faster doubling rate (p = 0.023); and PAS had a 4.7 hours shorter lag time (p = 0.016). CONCLUSION: We observed that five index organisms will grow in LR-AP stored in a 35%:65% ratio of plasma to InterSol where initial bacterial concentrations are 0.5 to 1.6 CFUs/mL. The more rapid initiation of log-phase growth for bacteria within a PAS storage environment resulted in a bacterial concentration up to 4 logs higher in the PAS units compared to the plasma units at 24 hours, but with no difference in the conc-max. This may present an early bacterial detection advantage for PAS-stored PLTs.


Assuntos
Plaquetas/microbiologia , Preservação de Sangue/métodos , Plaquetoferese , Infecções Estafilocócicas/sangue , Staphylococcus epidermidis/crescimento & desenvolvimento , Preservação de Sangue/efeitos adversos , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/sangue , Humanos , Infecções por Klebsiella/sangue , Klebsiella oxytoca/crescimento & desenvolvimento , Klebsiella oxytoca/isolamento & purificação , Plasma/microbiologia , Infecções por Serratia/sangue , Serratia marcescens/crescimento & desenvolvimento , Serratia marcescens/isolamento & purificação , Soluções , Staphylococcus aureus/crescimento & desenvolvimento , Staphylococcus aureus/isolamento & purificação , Staphylococcus epidermidis/isolamento & purificação , Infecções Estreptocócicas/sangue , Estreptococos Viridans/crescimento & desenvolvimento , Estreptococos Viridans/isolamento & purificação
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