electroporation transfer

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CAR-T Gene Packaging Delivery

• Reference https//www.creative-biolabs.com/car-t/
  gene-packaging-delivery.htm

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Background

• Since the successful manufacture of CAR-T cells
  is largely dependent on T cell receptor (TCR)
  gene transfer, this genetic modification of T
  lymphocytes is one of the most critical steps to
  generate superior CAR-T cells. Although the T
  cells are difficult to modify using non-viral
  methods like lipid-based car t plasmid
  transfection due to the high toxicity and low
  efficiency, the electroporation transfection and
  transposon based transfection systems are much
  more robust to complete the T-cell transfection.

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CAR-T cell transfection approaches
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Electroporation

• Electroporation is emerged as a powerful tool for
  the genetic modification of diverse cell types
  based on the transient disruption of cell
  membrane via exposure to an electric field, which
  allows charged molecules to enter the cell.
• For instance, the square-wave pulse-based new
  electroporation devices, such as Lonza
  Nucleofector II electroporation system, manifests
  a high efficiency in the genetic modification of
  T cells with proprietary electroporation buffers
  and electric parameters. In detail, up to 80 of
  viability and 40-60 of expression are achieved
  in human T cells.

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Retroviral transfection

• one of the mainstays of current gene therapy
  approaches, which contains a reverse
  transcriptase to enable the integration of
  artificial genes into the host genome in a stable
  status. Particularly, the retroviral vectors can
  either be replication-competent or
  replication-defective, and the latter is the most
  common choice for research since the viruses are
  competent for additional rounds of virion
  replication and packaging with other genes.
• With a typical maximum 8-10 kB length of an
  allowable DNA insertion in a replication-defective
  viral vector, the viruses are capable of
  infecting the target cells and delivering the
  specific viral payload. Meanwhile, the MLV fails
  to perform the typical lytic pathway and lead to
  cell lysis and death. Noticeably, MLV requires
  actively dividing cells for transduction.

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Lentiviral (complex retrovirus) transfection

• one of the mainstays of current gene therapy
  approaches, which enable the integration of
  artificial genes into the host genome in a stable
  status for genetical modifications in actively
  dividing cells as a subclass of retroviruses.
• More powerfully, the lentivirus has the ability
  to integrate foreign genes into the genome of
  non-dividing cells. Generally comparing to the
  electroporation, the virus-based transfections
  are relatively time-consuming and expensive,
  which limit a broader application in the clinical
  usage.

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Thanks

• Reference https//www.creative-biolabs.com/car-t/
  gene-packaging-delivery.htm