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A new promising CAR.CD30 T-cell therapy for CD30+ lymphoma.
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Figure 1. CAR.CD30 T cells with CD28.OX40 or CD28.4-1BB co- stimulation exhibit similar trans- duction levels and in vitro prolifer- ation upon cytokine stimulation. (A) Diagram of the expression cas- sette of two third-generation CAR.CD30. The single-chain vari- able fragment (scFv) of CD30 was cloned in frame with CD8aTM, CD28 cytoplasmic moiety, and a second co-stimulatory domain represented by either 4-1BB or OX40, as well as the signaling domain CD3-zeta chain (ζ). As a trackable marker, we added a peptide derived from human CD34 (DCD34). A safety switch, namely inducible caspase-9 (iCasp9), was also included in the vector constructs. (B) Flow-cytom- etry analyses in a representative donor showing chimeric antigen receptor (CAR) expression by detection of membrane CD34 in non-transduced (NT) T cells (nega- tive control; left panel), T cells genetically modified with CAR.CD30.ΔCD34.28.4.1BB.ζ (28.4.1BB.ζ; middle panel) and T cells genetically modified with CAR.CD30.ΔCD34.CD28.OX40.ζ (28.OX40.ζ; right panel). (C) Flow- cytometry analyses in a represen- tative donor showing CAR expres- sion using L protein to detect the scFv in NT T cells (negative con- trol; left panel), T cells genetically modified with 28.4.1BB.ζ (middle panel) and T cells genetically modified with 28.OX40.ζ (right panel). (D) Percentage of CAR+ CD3+ T cells during the time course of prolonged in vitro cul- ture (day +5 white bars; day +15 gray bars; day +30 black bars), in NT, 28.4.1BB.ζ and 28.OX40.ζ grown in interleukin (IL)2 or in IL7/IL15. (E, F) Fold expansion of NT (empty circle), 28.4.1BB.ζ (gray circle) and 28.OX40.ζ (black circle) grown in IL2 (E) or IL7/IL15 (F) (dotted lines). Data from seven healthy donors are expressed as average ± standard deviation. *P≤0.05; **P≤0.01; ***P≤0.001. Circled asterisks refer to the difference between the T-cell populations grown in IL2 or in IL7/IL15.
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haematologica | 2021; 106(4)
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