Evolution of CD19-Directed Chimeric Antigen Receptors (CARs) and Their Use in Clinical Trials Targeting Lymphoid Malignancies. All generations of CARs contain a transmembrane structural domain, as well as an extracellular single chain variable fragment (scFv), which is derived from a human-CD19-specific mouse monoclonal antibody — either FMC63 (IgG2a) or SJ25C1 (IgG1). First-generation CARs contain a single cytoplasmic signaling domain (CD3-ζ), which links antigen recognition to intracellular signal transduction pathways. Second-generation CARs contain CD3-ζ plus a co-stimulatory signaling domain, either CD28 or 4-1BB (also known as CD137, a member of the tumor necrosis factor receptor superfamily). Compared to first-generation CARs, second-generation CARs induce superior anti-tumor responses in preclinical studies and in patients with B-cell malignancies. Third-generation CARs contain two co-stimulatory domains, CD28 and 4-1BB, in addition to signaling domain CD3-ζ. Baylor (Baylor College of Medicine) – clinical trials NCT01853631, NCT00586391NCI (National Cancer Institute) – clinical trials NCT01087294, NCT00924326, NCT01593696MSKCC (Memorial Sloan-Kettering Cancer Center) – clinical trials NCT01840566, NCT01860937, NCT01044069, NCT00466531, NCT01416974Penn (Abramson Cancer Center of the University of Pennsylvania) – clinical trials NCT01747486, NCT01029366, NCT01551043