Table_1_T-Cell Immunotherapy for Pediatric High-Grade Gliomas: New Insights to Overcoming Therapeutic Challenges.xlsx

Despite decades of research, pediatric central nervous system (CNS) tumors remain the most debilitating, difficult to treat, and deadliest cancers. Current therapies, including radiation, chemotherapy, and/or surgery, are unable to cure these diseases and are associated with serious adverse effects and long-term impairments. Immunotherapy using chimeric antigen receptor (CAR) T cells has the potential to elucidate therapeutic antitumor immune responses that improve survival without the devastating adverse effects associated with other therapies. Yet, despite the outstanding performance of CAR T cells against hematologic malignancies, they have shown little success targeting brain tumors. This lack of efficacy is due to a scarcity of targetable antigens, interactions with the immune microenvironment, and physical and biological barriers limiting the homing and trafficking of CAR T cells to brain tumors. In this review, we summarize experiences with CAR T–cell therapy for pediatric CNS tumors in preclinical and clinical settings and focus on the current roadblocks and novel strategies to potentially overcome those therapeutic challenges.</p

Supplementary Figures from Inducible Activation of MyD88 and CD40 in CAR T Cells Results in Controllable and Potent Antitumor Activity in Preclinical Solid Tumor Models

Supplemental Figure 1: Activation of iCO molecules in T cells enhances NF-κB signaling;Supplemental Figure 2: Activation of iCO molecules enhances cytokine production in T cells;Supplemental Figure 3: CID dose-dependent IL2 production of HER2ζ.iCO T cells can be controlled by CID;Supplemental Figure 4: iCO activation enhances pro-inflammatory cytokine secretion of HER2ζ.iCO T cells;Supplemental Figure 5: Signal 1 is critical for HER2ζ.iCO T-cell effector function;Supplemental Figure 6: HER2ζ.iCO and HER2.CD28ζ T-cell lines have similar T-cell subset composition;Supplemental Figure 7: HER2ζ.iCO T cells have reduced PD-1 surface expression;Supplemental Figure 8: Recurrent tumors from HER2ζ.iCO T-cell + CID and HER2.CD28 ζ T-cell treated mice express HER2;Supplemental Figure 9: HER2ζ.iCO and HER2.CD28ζ T-cell lines have similar T-cell subset composition in vivo;Supplemental Figure 10: Anatomic location of tumors in treated mice;Supplemental Figure 11: Chemokine secretion of A549 cells, and chemokine receptor expression of T cells;Supplemental Figure 12: PD-L1 expression in tumor cell lines;Supplemental Figure 13: Generation of T-cells expressing HER2.CD28ζ.iCO;Supplemental Figure 14: iCO activation enhances expansion and pro-inflammatory cytokine secretion of HER2ζ.iCO and HER2.CD28ζ.iCO T-cells;Supplemental Figure 15: iCO activation enhances cytolytic activity of HER2ζ.iCO and HER2.CD28ζ.iCO T-cells; Supplemental Figure 16: Second CID injection eliminates recurrent tumors in 2/3 mice</p

Supplemental Table 1 from Transgenic Expression of IL15 Improves Antiglioma Activity of IL13Rα2-CAR T Cells but Results in Antigen Loss Variants

Tumor signal comparison.</p

Supplemental Figures 1 - 8 from Transgenic Expression of IL15 Improves Antiglioma Activity of IL13Rα2-CAR T Cells but Results in Antigen Loss Variants

Supplementary Figure 1: IL13Rα2-CAR and/or IL15 expression does not change T-cell phenotype. Supplementary Figure 2: Cell surface expression of IL13Rα2. Supplementary Figure 3: IL15 improves cell viability in the absence of exogenous cytokines. Supplementary Figure 4: Transgene expression is upregulated upon T cell activation. Supplemental Figure 5: IL13Rα2-CAR.IL15 T cells display activation-dependent IL15 production and greater proliferative capacity. Supplementary Figure 6: CID induces cell death in T cells genetically modified with retroviral vector encoding iC9.dNFGR.IL15. Supplemental Figure 7: Addition of exogenous IL15 increases CAR T-cell expansion in vitro. Supplementary Figure 8: IL13Rα2 or HER2 antigen loss variants are not killed by IL13Rα2- or HER2-CAR T cells.</p

Figures S1-S11 from Reversible Transgene Expression Reduces Fratricide and Permits 4-1BB Costimulation of CAR T Cells Directed to T-cell Malignancies

S1. Lack of functional exhaustion in expanding 28.z CD5 CAR T cells. S2. (A) Schematic representation of 28.z CD5 CARs with intermediate (CH3) and short spacer. (B) Expression level of CD5 CARs measured by flow cytometry using anti-mouse F(ab)-specific antibodies. (C) Expansion of T cells transduced with the 28.z CD5 CARs or BB.z CD5 CAR. Data are shown as mean and SD, n=3. S3. Relative frequency of CD45RA+ CCR7+ naïve-like (TNAIVE) and CD45RA- CCR7+ central memory (TCM) cells among T cells expressing indicated CD5 CAR constructs. Data are shown as mean and SD, n=3. S4. Reduced phosphorylation of IKKa/b in CD5 CAR T cells with mutant 4-1BB endodomain. S5. Relative frequency of CD45RA+ CCR7+ naïve-like (TNAIVE) and CD45RA- CCR7+ central memory (TCM) cells among T cells expressing indicated CD5 CAR constructs. Data are shown as mean and SD, n=3. S6. Morphology of T cells expressing indicated CAR constructs 5 days post-transduction. S7. (A) Expression of BB.z CD5 CAR in T cells transduced with retroviral (RV) or lentiviral (LV) vector. (B) Viability of BB.z CD5 CAR T cells 7 days post-transduction. (C) Expression of ICAM-1 on the surface of BB.z CD5 CAR T cells 7 days post-transduction. (D) In vivo control of systemic leukemia by 28.z and BB.z CD5 CAR T cells. (***, P<0.001 by one-way ANOVA with Bonferroni post-test correction). S8. Minimal CAR expression in the presence of DOX is insufficient to promote T-cell cytotoxicity. S9. Reciprocal expression of CD5 and CD5 CAR in Tet-OFF BB.z CD5 CAR T cells after DOX withdrawal. S10. Expansion (A), phenotype (B) and anti-tumor function of Tet-OFF 28.z CD5 CAR T cells in vitro (C). (D) CD5 CAR T cells were injected i.v. into mice with previously established systemic Jurkat leukemia (1x10^6 cells per mouse). Leukemia progression was monitored by IVIS imaging. S11. (A) CD5 CAR T cells with repressed (+DOX) or restored (-DOX) expression of BB.z CD5 CAR were injected i.v. into mice with previously established systemic Jurkat leukemia (1x10^6 cells per mouse). Leukemia progression was monitored by IVIS imaging. (B) Tumor levels in mice on day 34 post-tumor engraftment (*, P,0.05 by one-way ANOVA with Bonferroni post-test correction).</p