June 2022
The following articles have been recommended for further reading in the field of cancer immunotherapy by JITC's Immune Cell Therapies and Immune Cell Engineering Section Editor, Dr. Marcela Maus.
“Screening for CD19-specific chimeric antigen receptors with enhanced signaling via a barcoded library of intracellular domains"
by Khloe S. Gordon et al
Nature Biomedical Engineering (2022)
All current CAR T cells are constructed based on combinations of CD3z, CD28, and 4-1BB for intracellular immunostimulatory signaling. Khloe S. Gordon and colleagues pioneered a process termed CARPOOL to screen a library of 700,000 unique CD19-specific CARs with varied intracellular domains for enhanced anti-tumor properties. Jurkat cells transduced with barcoded CARs were selected for activation by CD69 and susceptibility to exhaustion by PD-1 over three rounds and then the ICDs and barcodes were long-read sequenced. Several novel intracellular domain combinations that have not been previously characterized that exhibited increased cytotoxicity and cytokine expression emerged from the screen. Some of the top hits from the screen were exhaustion-resistant against repeated antigen exposure and induced transcriptional programs consistent with lasting tumor memory. In mouse lymphoma models, the CARPOOL-identified CARs showed comparable tumor control to 4-1BB costimulated CAR T cells. One hit from the screen not only had activity against solid tumors, but led to sustained complete remissions—in contrast to 4-1BB costimulated CAR T cells that had a 100% relapse rate in the treated mice.
Why this matters: The CARPOOL method is a high-throughput strategy for identifying CARs with improved efficacy and persistence and may potentially be applied to developing cell therapies for tumor types that CAR T cells have been ineffective in treating thus far.
“Potentiating adoptive cell therapy using synthetic IL-9 receptors”
by Anusha Kalbasi et al
Nature (2022)
The efficacy of adoptive cell therapy is limited by poor in vivo proliferation and persistence of transferred T cells, necessitating conditioning with lymphodepleting chemotherapy. To reprogram adoptively transferred cells to a stem-like phenotype in vivo, Anusha Kalbasi et al designed a synthetic orthogonal cytokine-receptor pair. Orthogonal cytokines only bind to a mutant extracellular domain and do not cause signaling in tissues lacking the cognate orthogonal receptor. In this work, a previously generated orthogonal IL-2 extracellular domain was fused with the intracellular domain for IL-9 to generate the chimeric receptor o9R. In vitro, T cells carrying o9R phosphorylated STAT1, STAT3, and STAT5 and became phenotypically enriched for stem cell memory and effector markers upon stimulation with the orthogonal IL-2. In mice bearing B16 melanoma tumors, treatment with adoptively transferred gp100-specific T cells carrying the o9R receptor and orthogonal IL-2 led to anti-tumor efficacy even in the absence of lymphodepleting radiotherapy. Despite relatively low levels of proliferation, the o9R-expressing T cells infiltrated tumors and were enriched for activation markers and cytolytic capacity. Similarly high efficacy and enrichment for effector functions were observed in mice carrying mesothelin-expressing immunotherapy-resistant pancreatic ductal adenocarcinoma and treated with mesothelin-specific CAR T cells expressing o9R with orthogonal IL-2 delivered by an adenoviral vector. Notably, high rates of ICANS were observed with o9R-expressing CAR T cells, which may have reflected on-target/off-tumor activity driven by meningeal mesothelin expression. Like the murine models, human T cells transduced with a human orthogonal chimeric IL-9 receptor and an NY-ESO-1 cancer-testis antigen TCR demonstrated enrichment for stem and central memory phenotypes in culture upon stimulation with a human orthogonal IL-2 despite weaker proliferative signaling.
Why this matters: Orthogonal cytokine-receptor pairs represent a promising strategy to manipulate the phenotypes and persistence of adoptive cell therapy products in vivo, potentially streamlining manufacturing, reducing a need for lymphodepletion, and enhancing potency.
“A long-acting interleukin-7, rhIL-7-hyFc, enhances CAR T cell expansion, persistence, and anti-tumor activity”
by Yared Hailemichael et al
Nat Commun (2022)
Suboptimal CAR T cell efficacy often results in disease relapse even in the absence of antigen loss. Miriam Y. Kim et al demonstrate enhanced CAR T cell expansion and efficacy with in vivo stimulation with a recombinant long-lasting prolymphoid growth factor IL-7. Previously, rhIL-7-hyFc, a homodimeric IL-7 fused to a hybrid Fc platform to prevent complement activation was shown to be tolerable and increase CD4+ and CD8+ T cell counts in humans. Coculture with rhIL-7-hyFc led to significant expansion of CD19-specific CAR T cells while maintaining their functionality and enriching for central memory phenotype. In immunodeficient mice bearing aggressive human B cell lymphomas, treatment with combined 3rd generation CD28 and 4-1BB costimulated human anti-CD19 CAR T cell and rhIL-7-hyFc improved survival. Peripheral blood CAR T cells were expanded and skewed toward an effector memory phenotype in the mice. Similar antitumor efficacy was seen in immunodeficient mice engrafted with CD33+ acute myeloid leukemia and treated with a 2nd generation 4-1BB costimulated anti-CD33 CAR T cell plus rhIL7-hyFc. Notably, expansion of the anti-CD33 CAR T cells occurred only when administered with rhIL-7-HyFc. In immunocompetent models, co-administration of rhIL-7-hyFc to murine CAR T cells not only was associated with enhanced expansion and persistence without increased toxicity, but also reduced the effective dose. Furthermore, single cell transcriptomics revealed decreased exhaustion in peripheral CD4+ and CD8+ CAR T cells with rhIL-7-hyFc treatment.
Why this matters: Administration of rhIL-7-hyFc enhances not only the quantity but also the quality of CAR T cells, leading to better proliferation, survival, and efficacy. Given that rhIL-7-hyFc has been demonstrated to be safe in humans, this strategy warrants further investigation to prevent disease relapse.
“CD58 loss in tumor cells confers functional impairment of CAR T cells”
by Xin Yan et al
Blood Advances (2022)
CAR T cell therapy has improved outcomes for many patients with hematologic malignancies, however, a subset of patients have disease that do not respond to initial treatment. Using a CRISPR screening library targeting roughly 19,000 genes, Xin Yan and colleagues identified tumor genes involved primary resistance to CAR T cell therapy. CD58, a T cell activation co-stimulatory molecule, emerged as a top hit in the screen, which used CD19+ human pre-B cell acute lymphoblastic leukemia cells transduced with guide RNAs and cocultured with anti-CD19 CAR T cells for selection. The hit was validated in vitro, where disruption of the CD58/CD2 co-stimulatory axis in leukemia cells through genetic knockout or a blocking antibodies caused CAR T cells to become largely dysfunctional, with reduced expansion, degranulation, cytotoxicity, and secretion of cytokines such as IL-2, TNF-α, and IFN-y. Loss of CD58 prevented immune synapse formation with CAR T cells, resulting in reduced markers of activation and altered gene expression related to activation, proliferation, differentiation, and other important T cell functions. In a mouse xenograft model, tumors lacking CD58 were less efficiently cleared and survival was impaired with CAR T cell treatment compared to wild-type tumors with accompanying lower levels of CAR T cell peripheral expansion was reduced.
Why this matters: This study identifies the CD58/CD2 axis as an important mediator of CAR T cell functionality, providing rationale for future investigations of stimulation of the CD58/CD2—either pharmacologically or through novel CAR T constructs with CD2 co-stimulatory domains—to enhance response to CAR T cell therapy. These data may also inform studies of CD58 as a biomarker for resistance to CAR T cell therapy.