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Bidirectional Retroviral Integration Site PCR Methodology and Quantitative Data Analysis Workflow
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Longitudinal analysis of lentiviral and retroviral chimeric antigen receptors' integration sites reveals distinct

Vincent Guiraud1, Jerome Alexandre Denis2, Ghizlane Benhafoun1

  • 1Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Sante Publique (IPLESP), Assistance Publique-Hôpitaux de Paris (AP-HP), Hôpital Pitié-Salpêtrière, Service de Virologie, Paris, France.

British Journal of Haematology
|February 20, 2025
PubMed

Insights

Chimeric antigen receptor (CAR) T-cell therapy integration sites differ by vector type. Lentiviral CARs showed stable integration, while gammaretroviral CARs had variable persistence, necessitating further study for malignancy risk.

Area of Science:

  • Oncology
  • Immunotherapy
  • Molecular Biology

Background:

  • Chimeric antigen receptor (CAR) T-cell therapies offer potent cancer treatments.
  • CAR T-cell malignancies are linked to insertional mutagenesis, but longitudinal CAR integration site (IS) evolution is not well understood.

Purpose of the Study:

  • To investigate the longitudinal evolution of CAR integration sites (IS) following different CAR T-cell therapies.
  • To compare IS patterns and clonal persistence between lentiviral and gammaretroviral vectors.

Main Methods:

  • Integration site analysis in patient blood samples at peak CAR T-cell expansion and 1-year follow-up.
  • Analysis included patients treated with tisagenlecleucel (lentiviral), axicabtagene-ciloleucel, and brexucabtagene-autoleucel (gammaretrovirals).

Main Results:

  • Peak expansion IS patterns were vector-dependent: lentiviral CARs integrated in introns, while gammaretrovirals integrated in intergenic regions near transcription start sites.
  • One year post-infusion, lentiviral CAR showed no significant clonal proliferation.
  • Gammaretroviral CARs exhibited divergent outcomes: undetectable CAR in one patient and low-level oligoclonal persistence in another.

Conclusions:

  • CAR T-cell integration site patterns are influenced by vector type.
  • Longitudinal CAR T-cell persistence varies significantly between lentiviral and gammaretroviral vectors.
  • The long-term implications and potential malignancy risk associated with persistent gammaretroviral CARs require further investigation.
Keywords:
axicabtagene‐ciloleucelbrexucabtagene‐autoleucelchimeric antigen receptorclonal proliferationintegration sitetisagenlecleucel

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