Efficient Generation of Murine Chimeric Antigen Receptor (CAR)-T Cells

Rosa L. Vincent; Fangda Li; Edward R. Ballister; Nicholas Arpaia; Tal Danino

doi:10.3791/65887

Generación eficiente de células T con receptor de antígeno quimérico (CAR) murino

JoVE Journal
Bioengineering

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JoVE Journal Bioengineering

Efficient Generation of Murine Chimeric Antigen Receptor (CAR)-T Cells

Generación eficiente de células T con receptor de antígeno quimérico (CAR) murino

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06:22 min

February 2, 2024

DOI: 10.3791/65887-v

Rosa L. Vincent¹, Fangda Li², Edward R. Ballister¹, Nicholas Arpaia^2,3, Tal Danino^1,3,4

¹Department of Biomedical Engineering,Columbia University, ²Department of Microbiology & Immunology, Vagelos College of Physicians and Surgeons,Columbia University, ³Herbert Irving Comprehensive Cancer Center,Columbia University, ⁴Data Science Institute,Columbia University

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Overview

This protocol streamlines retroviral vector production and murine T cell transduction, facilitating the efficient generation of mouse CAR-T cells. The study focuses on the integration of engineered bacteria and CAR-T cells for solid tumor treatment in syngeneic models.

Key Study Components

Area of Science

Neuroscience
Immunology
Oncology

Background

CAR-T cell therapy is a promising approach for cancer treatment.
Murine models are essential for studying CAR-T cell efficacy.
Standardized protocols for generating murine CAR-T cells are lacking.
Transduction of murine T-cells is challenging with traditional methods.

Purpose of Study

To develop a streamlined protocol for murine CAR-T cell production.
To enable effective study of CAR-T cells in syngeneic tumor models.
To evaluate the activity of new CAR technologies in a complete immune system context.

Methods Used

Preparation of reduced serum medium for transduction.
Dilution of media with transfection reagent.
Application of the protocol to murine T cells.
Evaluation of CAR-T cell functionality in syngeneic models.

Main Results

The protocol successfully enhances murine T cell transduction efficiency.
Generated CAR-T cells exhibit improved functionality in tumor models.
Standardization allows for reproducibility in research.
Facilitates further exploration of CAR technologies.

Conclusions

This protocol provides a reliable method for generating murine CAR-T cells.
It supports the study of CAR-T cell interactions within the immune system.
Future research can build on this foundation to enhance cancer therapies.

Frequently Asked Questions

What are CAR-T cells?

CAR-T cells are genetically engineered T cells designed to target and kill cancer cells.

Why are murine models used in this research?

Murine models provide a relevant biological context to study immune responses and cancer therapies.

What challenges exist in murine T cell transduction?

Murine T cells are often difficult to transduce and culture using traditional methods.

How does this protocol improve CAR-T cell production?

The protocol streamlines the production process, enhancing transduction efficiency and reproducibility.

What implications does this research have for cancer treatment?

It paves the way for improved CAR-T cell therapies and better understanding of their mechanisms in solid tumors.

Can this protocol be adapted for other types of cells?

While designed for murine T cells, the principles may be applicable to other cell types with modifications.

Este protocolo agiliza la producción de vectores retrovirales y la transducción de células T murinas, lo que facilita la generación eficiente de células CAR-T de ratón.

Nuestra investigación explora combinaciones efectivas de bacterias modificadas genéticamente y células T con CAR para el tratamiento de tumores sólidos. Es importante para nosotros estudiar cómo funcionan estas dos terapias celulares en el contexto de un sistema inmunitario completo utilizando modelos singénicos de cáncer de ovario. El estudio de la función de las células T con CAR en modelos tumorales singénicos se ha visto limitado por la falta de protocolos estandarizados para generar células T con CAR murinas.

Las células T murinas son particularmente difíciles de transducir en cultivo ex vivo utilizando métodos tradicionales. Desarrollamos este protocolo para agilizar la producción de células CAR-T murinas para permitir el estudio en modelos singénicos que proporcionan un contexto importante para evaluar la actividad de las nuevas tecnologías CAR.

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