RESUMO
Chimeric antigen receptor (CAR) T-cell therapy is a breakthrough in hematologic malignancies, such as acute B lymphoblastic leukemia (B-ALL). Monitoring this treatment is recommended, although standardized protocols have not been developed yet. This work compares two flow cytometry monitoring strategies and correlates this technique with qPCR method. CAR-T cells were detected by two different flow-cytometry protocols (A and B) in nine blood samples from one healthy donor and five B-ALL patients treated with Tisagenlecleucel (Kymriah®, USA). HIV-1 viral load allowed CAR detection by qPCR, using samples from seven healthy donors and nine B-ALL patients. CAR detection by protocol A and B did not yield statistically significant differences (1.9% vs. 11.8% CD3 + CAR+, p = 0.07). However, protocol B showed a better discrimination of the CD3 + CAR+ population. A strong correlation was observed between protocol B and qPCR (r = 0.7, p < 0.0001). CD3 + CAR+ cells were detected by flow cytometry only when HIV-1 viral load was above 104 copies/mL. In conclusion, protocol B was the most specific flow-cytometry procedure for the identification of CAR-T cells and showed a high correlation with qPCR. Further efforts are needed to achieve a standardized monitoring approach.
Assuntos
Citometria de Fluxo , HIV-1 , Imunoterapia Adotiva , Receptores de Antígenos Quiméricos , Linfócitos T , Carga Viral , Humanos , Citometria de Fluxo/métodos , Imunoterapia Adotiva/métodos , HIV-1/imunologia , HIV-1/genética , Carga Viral/métodos , Receptores de Antígenos Quiméricos/imunologia , Linfócitos T/imunologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase em Tempo Real/normas , Complexo CD3 , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologiaRESUMO
Autoimmune lymphoproliferative syndrome is a primary immunodeficiency caused by variants in FAS-mediated apoptosis related genes and is characterized by lymphadenopathy, splenomegaly and autoimmunity. A total of six different variants in CASP10 have been described as potential causative of disease, although two of them have recently been considered polymorphisms. The high allele frequency of these variants in healthy population in addition to the broad clinical spectrum of the disease difficult the interpretation of their pathogenicity. Here, we describe the clinical and analytical findings of three new patients carrying variants in CASP10 and summarize 12 more cases from the literature. Autoimmune cytopenias, adenopathies and increment of TCRαß+CD4-CD8- cells have been the most common findings, being possibly the FAS-mediated apoptosis pathway the pathogenic mechanism of this disease. The clinical impact and the consequences of CASP10 variants are not fully elucidated, therefore the description of new cases will contribute to solve this issue.