Main description:

This volume provides comprehensive methods from expert scientists working in the Chimeric Antigen Receptor T Cell (CAR-T Cell) field. Chapters guide readers through the state-of-art of CAR-T cell technology, CAR design and vector production, CAR-T cell generation and manufacturing, CAR-T cell characterization, and quality control. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Authoritative and cutting-edge, Chimeric Antigen Receptor T Cells: Development and Production aims to be useful in the production of CAR-T cells, especially for therapeutic purposes.

Contents:

1. CAR-T Cells for Cancer Treatment: Current Design and Next Frontiers

Virginia Picanco-Castro, Kamilla Swiech, Kelen Cristina Ribeiro Malmegrim de Farias, and Dimas Tadeu Covas

Part I: Design of Chimeric Antigen Receptors and Vector Production

2. Selection of Antibody Fragments for CAR-T Cell Therapy from Phage Display Libraries

Nestor F. Leyton-Vastro, Marcelo M. Brigido, and Andrea Q. Maranhao

3. Phase I/II Manufacture of Lentiviral Vectors under GMP in an Academic Setting

Anindya Dasgupta, Stuart Tinch, Kathleen Szczur, Rebecca Ernst, Nathaniel Shryock, Courtney Kaylor, Kendall Lewis, Eric Day, Timmy Truong, and William Swaney

4. Production of Lentivirus for the Establishment of CAR-T Cells

Marlous G. Lana and Bryan E. Strauss

5. Optimized Production of Lentiviral Vectors for CAR-T Cell

Pablo Diego Moco, Mario Soares de Abreu Neto, Daianne Maciely Carvalho Fantacini, and Virginia Picanco-Castro

6. Lentiviral Vectors Production in Suspension Culture Using Serum-Free Medium for the Transduction of CAR-T Cells

Aline Do Minh, Michelle Yen Tran, and Amine A. Kamen

Part II: CAR-T Cells Generation and Manufacturing

7. In vitro Transcribed (IVT)-mRNA CAR Therapy Development

Miliotou N. Androulla and Papadopoulou C. Lefkothea

8. Generation of Chimeric Antigen Receptor T Cells Using Gammaretroviral Vectors

Feiyan Mo and Maksim Mamonkin

9. Generation of CAR+ T Lymphocytes using the Sleeping Beauty Transposon System

Leonardo Chicaybam, Luiza Abdo, and Martin H. Bonamino

10. Platforms for Clinical-grade Car-T Cell Expansion

Amanda Mizukami and Kamilla Swiech

11. CAR-T Cell Expansion in a Xuri Cell Expansion System W25

Trevor A. Smith

12. Methods and Process Optimization for Large Scale CAR T Expansion Using the G-Rex Cell Culture Platform

Josh Ludwig and Mark Hirschel

Part III: CAR-T Cells Characterization and Quality Control

13. Replication Competent Lentivirus Analysis of Vector Transduced T Cell Products used in Cancer Immunotherapy Clinical Trials

Kenneth Cornetta, Sue Koop, Emily Nance, Kimberley House, and Lisa Duffy

14. Immunophenotypic Analysis of CAR-T Cells

Julia Teixeira Cottas de Azevedo, Amanda Mizukami, Pablo Diego Moco and Kelen Cristina Ribeiro Malmegrim

15. Approaches of T Cell Activation and Differentiation for CAR-T Cell Therapies

Robert D. Schwab, Darel Martinez Bedoya, Tiffany R. King, Bruce L. Levine, and Avery D. Posey, Jr.

16. Determination of Cytotoxic Potential of CAR-T Cells in Co-cultivation Assays

Renata Silvestre Nacasaki, Pablo Diego Moco, and Virginia Picanco-Castro

17. Analysis of CAR-mediated Tonic Signaling

Hugo Calderon, Maksim Mamonkin, and Sonia Guedan

18. Generation of Tumor Cells Expressing Firefly Luciferase (fLuc) to Evaluate the Effectiveness of CAR in a Murine Model

Marcelo de Souza Fernandes Pereira, Daianne Maciely Carvalho Fantacini, and Virginia Picanco e Castro

19. Analysis of Antitumor Effects of CAR-T Cells in Mice with Solid Tumors

Alba Rodriguez-Garcia, Keisuke Watanabe, and Sonia Guedan