A Real-Time Quantitative PCR Targeting the Viral Vector for the Monitoring of Patients Treated with Axicabtagene Ciloleucel
| Autor: | L. Bocket, Pauline Varlet, Ibrahim Yakoub-Agha, Didier Hober, Agathe Baras, Enagnon Kazali Alidjinou, Christophe Hallaert, David Beauvais |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Male T cell Genetic Vectors Biology Real-Time Polymerase Chain Reaction Immunotherapy Adoptive Sensitivity and Specificity CD19 Virus Pathology and Forensic Medicine Viral vector Flow cytometry 03 medical and health sciences 0302 clinical medicine Antineoplastic Agents Immunological medicine Humans Digital polymerase chain reaction Transgenes Aged Monitoring Physiologic Biological Products Receptors Chimeric Antigen medicine.diagnostic_test Middle Aged Molecular biology Chimeric antigen receptor Data Accuracy 030104 developmental biology medicine.anatomical_structure Real-time polymerase chain reaction Treatment Outcome 030220 oncology & carcinogenesis biology.protein Molecular Medicine Female Lymphoma Large B-Cell Diffuse Gammaretrovirus |
| Zdroj: | The Journal of molecular diagnostics : JMD. 23(4) |
| ISSN: | 1943-7811 |
| Popis: | Axicabtagene ciloleucel or axi-cel [CD19 chimeric antigen receptor (CAR) T cell] has been recently approved for refractory/relapsed diffuse large B-cell lymphoma and primary mediastinal B-cell lymphoma. Proliferation of CAR T cells after infusion and their persistence have been reported as important factors. Laboratory tools are needed for the monitoring of patients. We developed a vector-based, simple, and accurate real-time quantitative PCR (qPCR) to measure axi-cel vector copy number in peripheral blood samples. Primers and probe targeting the 5′LTR region of the gammaretroviral vector (mouse stem cell virus) were designed for amplification. To generate standard curves, mouse stem cell virus plasmid was subcultured and quantified using droplet digital PCR. The method was applied to quantify vector copy number in blood samples from patients treated with axi-cel. The limit of quantification of the qPCR assay was established at 2.2 copies/μL in DNA eluate. The qPCR method was well correlated with flow cytometry findings; however, the assay appeared to be more sensitive than flow cytometry. The kinetics observed in blood samples from treated patients were in agreement with previously reported findings. In conclusion, we developed a sensitive and accurate qPCR assay for the quantification of transgenic CAR T cells, which can be a useful additional tool for the monitoring of patients treated with axi-cel. |
| Databáze: | OpenAIRE |
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