A Long-Acting Pharmacological Grade Interleukin-7 Molecule Logarithmically Accelerates CART Proliferation, Differentiation, and Tumor Killing

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Background CART therapy is revolutionizing modern cancer therapy, with two anti-CD19 CARTs FDA-approved for relapsed/refractory B cell lymphoma/leukemia and many other CARTs for solid and liquid tumors currently undergoing clinical trials. However, in current clinical practice, suboptimal CART persistence and tumor killing permit tumor cell escape and, ultimately, disease relapse. Reasoning that a pro-lymphoid growth factor could promote CART efficacy, we supplemented UCART infusion with subcutaneous injections of the long-acting form of recombinant human interleukin-7 fused with hybrid Fc (rhIL-7-hyFc, NT-I7) in vivo using a CD19+ lymphoma xenograft model. Methods To create anti-CD19 universal CART (UCART19), we activated human T cells on CD3/CD28 beads, electroporated the T cells with Cas9 mRNA and a TRAC gRNA, and virally transduced an anti-CD19 scFv 3rd generation CAR containing a peptidase 2A-cleaved human CD34 construct for both purification and tracking in vivo. Residual TRAC+ cells were depleted using magnetic selection. For xenograft tumor modeling in vivo, we injected NSG mice with 5 × 105 RamosCBR-GFP cells four days prior to UCART19 (2 × 106 cells). Mice were treated with NT-I7 (10mg/kg SC) on days +1, +15 and +29 post UCART19 infusion. Results RamosCBR-GFP mice receiving NT-I7 without UCART19 (NT-I7 only group) survived marginally longer (24 day med survival) than mice receiving RamosCBR-GFP cells alone (No tx group) (21 day medium survival, p=0.018, NT-I7 only vs. No Tx). While RamosCBR-GFP mice treated with UCART19 alone (UCART19 group) survived 33 days, 100% of RamosCBR-GFP mice treated with UCART19 and NT-I7 (UCART19+NT-I7 group) were alive at 80 days (Fig 1a), with no mouse showing signs of xenogeneic GVHD (p Discussion Here, we demonstrate that a pharmacological grade long-acting interleukin-7 agonist can potentiate adoptive cellular therapies. Specifically, NT-I7 can dramatically enhance gene modified T cell proliferation, persistence and tumor killing in vivo, resulting in enhanced survival. Unlike genetic strategies to potentiate IL-7 signaling in CART, NT-I7 provides a tunable clinic-ready adjuvant for reversing suboptimal CART activity in vivo.