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01.12.2018 | Research | Ausgabe 1/2018 Open Access

Journal of Hematology & Oncology 1/2018

CAR-T cells targeting CLL-1 as an approach to treat acute myeloid leukemia

Zeitschrift:
Journal of Hematology & Oncology > Ausgabe 1/2018
Autoren:
Jinghua Wang, Siyu Chen, Wei Xiao, Wende Li, Liang Wang, Shuo Yang, Weida Wang, Liping Xu, Shuangye Liao, Wenjian Liu, Yang Wang, Nawei Liu, Jianeng Zhang, Xiaojun Xia, Tiebang Kang, Gong Chen, Xiuyu Cai, Han Yang, Xing Zhang, Yue Lu, Penghui Zhou
Wichtige Hinweise

Electronic supplementary material

The online version of this article (https://​doi.​org/​10.​1186/​s13045-017-0553-5) contains supplementary material, which is available to authorized users.
Jinghua Wang, Siyu Chen and Wei Xiao are co-first authors.
Xing Zhang, Yue Lu and Penghui Zhou are co-last authors.

Abstract

Background

Acute myeloid leukemia (AML) is one of the most common types of adult acute leukemia. Standard chemotherapies can induce complete remission in selected patients; however, a majority of patients eventually relapse and succumb to the disease. Thus, the development of novel therapeutics for AML is urgently needed. Human C-type lectin-like molecule-1 (CLL-1) is a type II transmembrane glycoprotein, and its expression is restricted to myeloid cells and the majority of AML blasts. Moreover, CLL-1 is expressed in leukemia stem cells (LSCs), but absent in hematopoietic stem cells (HSCs), which may provide a potential therapeutic target for AML treatment.

Methods

We tested the expression of CLL-1 antigen on peripheral blood cells and bone marrow cells in healthy donor and AML patients. Then, we developed a chimeric antigen receptor (CAR) containing a CLL1-specific single-chain variable fragment, in combination with CD28, 4-1BB costimulatory domains, and CD3-ζ signaling domain. We further investigate the function of CLL-1 CAR-T cells.

Results

The CLL-1 CAR-T cells specifically lysed CLL-1+ cell lines as well as primary AML patient samples in vitro. Strong anti-leukemic activity was observed in vivo by using a xenograft model of disseminated AML. Importantly, CLL-1+ myeloid progenitor cells and mature myeloid cells were specifically eliminated by CLL-1 CAR-T cells, while normal HSCs were not targeted due to the lack of CLL-1 expression.

Conclusions

CLL-1 CAR-T represents a promising immunotherapy for the treatment of AML.

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Zusatzmaterial
Additional file 1: Table S1. Patient characteristics and CLL-1 expression of primary AML patient sample. F, female; M, male; BM, bone marrow; PB, peripheral blood. (DOCX 25 kb)
13045_2017_553_MOESM1_ESM.docx
Additional file 2: Figure S1. Co-expression CLL-1 and CD33 in primary AML samples. (A) Initially, cells were gated based on forward and side scatter properties. Subsequently, AML blasts were selected based on low side scatter versus CD45dim expression. (B) CLL-1 and CD33 expression on four representative gated AML blast cell populations are depicted. Percentages in each quadrant are indicated. (TIFF 1776 kb)
13045_2017_553_MOESM2_ESM.tif
Additional file 3: Figure S2. The gating strategy of CD34+ AML blasts. Cells were initially gated based on forward and side scatter properties. Subsequently, AML blasts were selected based on low side scatter versus CD45dim expression. Then, CD34+ cells were gated. Finally, CD38+/CD38− cells were gated and used for CLL-1 expression analysis. (TIFF 1656 kb)
13045_2017_553_MOESM3_ESM.tif
Additional file 4: Figure S3. CLL-1 CAR-T cells lyse CLL1-expressing AML cells. (A) Expression of CLL-1 on the cell lines HL-60 and K562. (B) CLL-1 CAR-T cells lysed CLL-1+ cell line HL-60. CLL-1− cell line K562 was used as negative control. NT cells were used to evaluate unspecific lysis. Data represent mean values of triplicate wells ± SD. (TIFF 481 kb)
13045_2017_553_MOESM4_ESM.tif
Additional file 5: Figure S4. Proliferation of CLL-1 CAR-T cells in response to CLL-1+ cells. Pair-matched CFSE-labeled CLL-1 CAR-T cells or NT cells were co-cultured with the indicated stimulator cell lines for 96 h at an E:T of 1:1. CFSE dilution was analyzed by flow cytometry. Unstimulated T cells (gray histograms) were used as baseline T cell proliferation controls. (TIFF 866 kb)
13045_2017_553_MOESM5_ESM.tif
Additional file 6: Figure S5. CLL-1 CAR expression in T cells derived from AML patients. T cells from three AML patients were transduced with CLL-1 CAR. Shown are CLL-1 CAR-T and NT cells from the three AML patients 14 days post transduction. Percentages in each quadrant are indicated. (TIFF 557 kb)
13045_2017_553_MOESM6_ESM.tif
Additional file 7: Figure S6. Representative flow cytometric analysis of peripheral blood of CAR-T-treated mice. Eighteen days after leukemia transplant, hCD45+ CLL1− population in peripheral blood of CAR-T-treated mice was almost human T cells (hCD45+ CD3+). (TIFF 4614 kb)
13045_2017_553_MOESM7_ESM.tif
Literatur
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