CAR T-cell therapy is a type of adoptive cell therapy to treat cancer. Using genetic engineering, T cells from patients are equipped with chimeric antigen receptor (CAR) to specifically recognize tumor antigen and kill tumor cells. However, CAR-T cell therapy is facing major clinical challenges including cytokine release syndrome and poor persistence. Therefore, it is important to design CAR-T cells with high persistence, little side effects and strong anti-tumor activity.
Recently, Dr. Chenqi Xu’s group at CAS Center for Excellence in Molecular Cell Science, Dr. Catherine Chiulan Wong’s group at Peking University and Dr. Enfu Hui’s group at University California, San Diego developed a new strategy of CAR-T cell therapy, which has reduced cytokine production, enhanced cell persistence and better anti-tumor function than clinical used 28Z CAR-T cells. This study was published in Cell entitled “Multiple Signaling Roles of CD3ε and Its Application in CAR-T Cell Therapy” on July 29.
T cell receptor (TCR) mediates antigen-induced signaling through its associated CD3ε, δ, γ, and ζ. Using quantitative mass spectrometry, the authors simultaneously quantitated the phosphorylation of the immunoreceptor tyrosine-based activation motif (ITAM) of all CD3 chains upon TCR stimulation. The results showed that CD3ε ITAMs was mono-phosphorylated, owing to Lck kinase selectivity, and specifically recruited the inhibitory Csk kinase to attenuate TCR signaling, suggesting that TCR is a self-restrained signaling machinery containing both activating and inhibitory motifs. Next, the authors incorporated CD3ε cytoplasmic domain into the clinical used 28Z CAR, and found that the new E28Z CAR-T cells had improved antitumor activity. Mechanistically, the Csk-recruiting ITAM of CD3ε reduced CAR-T cell cytokine production whereas the basic residue rich sequence (BRS) of CD3ε promoted CAR-T cell persistence via p85 recruitment. Considering the strong anti-tumor activity of E28Z CAR-T cells in both blood tumor and solid tumor models, these results provide a solid rationale to explore its promising translational potentials in treating blood and solid malignancies.
Ph.D. candidates Wei Wu, Qiuping Zhou and Xiaoshan Shi from CAS Center for Excellence in Molecular Cell Science and Dr. Takeya Masubuchi from University California, San Diego are the co-first authors. Dr. Chenqi Xu, Dr. Catherine Chiulan Wong and Dr. Enfu Hui are the co-corresponding authors. Dr. Haopeng Wang from ShanghaiTech University and Dr. Jie Sun from Zhejiang University participate the study. Dr. Chenqi Xu is the adjunct researcher of Hangzhou Institute For Advanced Study of the Chinese Academy of Sciences.
Clinical used 28Z CAR-T cells are facing major clinical challenges including cytokine release syndrome and poor persistence. E28Z CAR-T cells incorporating the CD3ε cytoplasmic domain have reduced cytokine production due to Csk-recruiting ITAM of CD3εand enhanced persistence through p85- recruiting BRS of CD3ε.
T cell therapies have achieved great success in treating hematologic malignancies but facing difficulties in treating solid tumor. Chenqi Xu was invited by Cancer Cell to contribute a preview discussing a CRISPR-based platform for developing T cell therapies, which allows discovery of constructs promoting T cell anti-tumor activity.
In a recent Cell paper by Marson and colleagues, the researchers developed a method for pooled knockin screening of large DNA sequences at the endogenous TCRa constant (TRAC) locus. They designed a 36-member library of barcoded templates, which enables the quantification of on-target integration of each construct. They further developed pooled knockin sequencing (PoKI-seq), combing single-cell transcriptome analysis and pooled knockin screening to measure cell abundance and cell states ex vivo and in vivo. Using this innovative technology, they found that a novel chimeric TGF-bR2-41BB receptor, which convert suppressive TGFb signaling to stimulatory 41BB signaling, enhanced T cell fitness and thus promoted solid tumor clearance. This pooled knockin platform will accelerate discovery of new constructs for T cell therapies.
PD-1, a well-established drug target for cancer immunotherapy, has been recognized as a driver of T cell exhaustion for long time but this concept is now challenged by a recently published paper at Molecular Cell by Okazaki and colleagues. Chenqi Xu was invited by Molecular Cell to contribute a preview discussing these controversial data and proposing his own model of PD-1 function.
In the paper of Okazaki and colleagues, the authors used human and mouse T cell systems to study how PD-1 signaling impacts transcriptome at the early stage of T cell activation. They find PD-1 primarily affects genes that are induced or suppressed by strong TCR signaling, i.e. cytokine genes but not proliferation and exhaustion genes. This paper points out PD-1’s role at the early stage of T cell activation is controlling effector function but not inducing exhaustion. One needs to notice that PD-1 expression is only transient during normal T cell activation. Therefore, the new paper more reflects the function of transient PD-1 signaling. However, at the contexts of cancer or chronic infection, PD-1 expression is persistently induced by chronic antigen exposure. Persistent PD-1 signaling can work together with key transcription factors to induce and reinforce T cell exhaustion status. In summary, PD-1’s role is different under different scenarios and its contribution to T cell exhaustion needs to be further studied.
Temporarily Free link: https://authors.elsevier.com/a/1agzW3vVUPDe88
The T cell receptor (TCR) is one of the most complicated receptors in mammalian cells. Attracted by the complexity and functional importance of TCR, many groups have been studying TCR structure and triggering for decades using diverse biochemical and biophysical tools. Recently, a review, entitled “structural understanding of T cell receptor triggering”, by the research group led by Dr. Chenqi Xu at Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, is published online at Cellular & Molecular Immunology. The review synthesizes the structural studies and discusses the relevance of the conformational change model in TCR triggering.
T cells play essential roles in the adaptive immune response against pathogens and cancer cells. T cells specifically recognize peptide antigens loaded on the major histocompatibility complex (peptide-MHC, pMHC) upon TCR activation. As an octamer complex, TCR comprises an antigen-binding subunit (TCRαβ) and three CD3 signaling subunits (CD3γε, CD3δε, and CD3ζζ). Engagement of TCRαβ with an antigen pMHC leads to tyrosine phosphorylation of the immunoreceptor tyrosine-based activation motif (ITAM) in CD3 cytoplasmic domains, thus translating extracellular binding kinetics to intracellular signaling events. To explain the triggering mechanism of TCR, several models have been proposed, including but not limited to kinetic segregation, serial engagement, kinetic proofreading, and conformational change. Recently, a signiﬁcant breakthrough was made in the TCR ﬁeld. Dong et al. reported a cryo-electron microscopy (cryo-EM) structure of a human TCR–CD3 complex in its unliganded state at 3.7 Å resolution (Nature. 573 (7775): 546-552, 2019). As expected, the TCR-CD3 complex is assembled with a 1:1:1:1 stoichiometry of TCRαβ/ CD3γε/CD3δε/CD3ζζ. Whether conformational change plays an important role in the transmembrane signal transduction of TCR is thus brought into attention again. In the review, we highlight the conformational changes that occur in the extracellular, transmembrane, and intracellular domains upon TCR triggering. Most results corroborate each other and are useful for revealing the mechanism of TCR triggering. Since lipids play essential roles in regulating TCR structure and function, studying the TCR-CD3 complex in a native membrane environment is warranted in future research. Moreover, T cell-based immunotherapies such as TCR-T and CAR-T can be further developed and applied in the clinic based on a better understanding of TCR triggering.
Prof. Chenqi Xu serves as the corresponding author to design the framework and extensively revise the manuscript. Xinyi Xu and Hua Li, a graduate student and an associate professor in the Xu Lab, serve as co-first authors. The work was supported by grants from NSFC grants, CAS grants (Strategic Priority Research Program; Facility-based Open Research Program; Fountain-Valley Life Sciences Fund of University of Chinese Academy of Sciences Education Foundation), MOST, and the Ten Thousand Talent Program “Leading Talent” of China.
Paper link: https://rdcu.be/b1wik.