Additional file 1 of Expanding anti-CD38 immunotherapy for lymphoid malignancies
Creators: name: Wang, Xu
name: Yu, Xinfang
name: Li, Wei
name: Neeli, Praveen
name: Liu, Ming
name: Li, Ling
name: Zhang, Mingzhi
name: Fang, Xiaosheng
name: Young, Ken H.
name: Li, Yong
Publisher: name: figshare
Year: 2022
Subjects: subject: Immunology
subject: FOS: Clinical medicine

description: Additionalfile 1: Supplemental Figure S1. Anti-myelomafunction of CD38-CAR T cells in vitro. (A) CD38 expression in MM cell lines (MM.1S, NCI-H929, OPM-1, RPMI-8226, KMS-12, and ANBL-6) measured by flow cytometry. (B) Thecytotoxic activity of CD4+ and CD8+ CD38-CAR T cells. T cells were co-cultured with MM cells and K562 (negative control) with stably expressed fLuc for 4 and 20 h, respectively, at various ratios of E:T. (C) Evaluation of CD107a expression by flow cytometry. Effector cells were co-cultured with target cells for 6 h at 2:1 E:T ratio. (D) Proliferation assessed by absolute cell number and CellTrace™ far-red proliferation dilution after a 5-day co-culture of effector and target cells. Assays were performed with effector cells and irradiated target cells at 1:2 E:T ratio without the addition of exogenous cytokines. (E)Secretion of IL-2, IFN-γ, TNF-α, and perforin from effector cells by ELISA. Assays were performed in supernatants obtained after a 20-h co-culture of effector and target cells at a 2:1 E:T ratio. Supplemental Figure S2. Anti-myeloma function of CD38-CAR T cells invivo. (A) Schematic of CAR T cells treatment protocol. NSG mice were intravenously injected with 1 × 106 fLuc-transduced MM cells (OPM-1). After 7 days, the mice were treated with 5×106 of CD38-CAR T cells, control T cells, or remained untreated (n = 5 or 4). (B) A serial of representative bioluminescent imaging showing myeloma progression/regression in mice. (C) Kinetics of myeloma progression in mice treated with CD38-CAR T cells, control T cells, or remained untreated measured by bioluminescent imaging. (D) Survival curve for each experimental group. Supplemental Figure S3. CD38-CAR T cells inhibit NKTCL growth in the prophylactic xenograftmodel. (A) Schematic of CAR T cells treatment protocol. NSG mice were subcutaneously injected with 5×106 NKTCL cells (YT). After 10 days, the mice weretreated with 5×106 of CD38-CAR T cells or control T cells (n= 5). (B) Mean tumor growth kinetics for each treatment group. Supplemental FigureS4. ATRA enhances the CD38 expression in MM, MCL, and DLBCL. (A–C) CD38 expression in MM, MCL, or DLBCL with or without ATRA (10 nM) treatment by flow cytometry. (D) CD27 expression in MCL cell lines with or without ATRA (10 nM) treatment (JeKo-1,Granta-519, Mino and SP-53) by flow cytometry. (E) Mino and Granta-519 were treated with ATRA (10 nM) or PF-431396 for 48 hours. CD38 expression were evaluated by immunoblotting. Supplemental Figure S5. Retinoids enhance the CD38 expression invariable degrees. (A) SP-53 cells were treated with adapalene, tazarotene,acitretin, isotretinoin, or the vehicle control ranging from 10 to 200 nM for 24 or 48 h. (B) The fold change of median fluorescence intensity (MFI) of CD38 in SP-53 cells after treatment of ATRA (10 nM) or four other retinoids for 24 or 48 h. Supplemental Figure S6. Hematoxylin and eosin (H&amp;E) and immunohistochemistry (IHC) staining for tumors from mice with ATRA or anti-CD38-based immunotherapy. (A) Representative IHC images of indicated treatment SP-53 tumors for CD38. Scalebars, 100 μm; H&amp;E staining of SP-53 tumors on day 22. Scale bars, 100 μm. (B) Representative IHC images of indicated treatment SP-53 tumors for CD38. Scale bars, 100 μm; H&amp;E staining of SP-53 tumors on day 22. Scale bars, 100μm. Supplemental Table 1. Cell lines used in this study.