2024年10月，中國科學(xué)院動(dòng)物研究所干細(xì)胞與生殖生物學(xué)國家重點(diǎn)實(shí)驗(yàn)室；北京干細(xì)胞與再生醫(yī)學(xué)研究所；暨南大學(xué)第一附屬醫(yī)院血液科(State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China；Beijing Institute for Stem Cell and Regenerative Medicine, Beijing, China；Department of Hematology, The First Affiliated Hospital of Jinan University, Guangzhou, China) Qi Zhang老師研究團(tuán)隊(duì)在《Cancer Immunity and Immunotherapy》上發(fā)表論文：

“Hypoimmunogenic CD19 CAR-NK cells derived from embryonic stem cells suppress the progression of human B-cell malignancies in xenograft animals"

“來自胚胎干細(xì)胞的低免疫原性CD19 CAR-NK細(xì)胞抑制異種移植動(dòng)物中人類b細(xì)胞惡性腫瘤的進(jìn)展"

Abstract：

Background: Chimeric antigen receptor (CAR) engineered natural killer (NK) cells exhibit advantages such as MHC-independent recognition and strong anti-tumor functions. However, allogeneic CAR-NK cells derived from human tissues are heterogeneous and susceptible to clearance by hosts.

Methods: We generated a B2M knockout, HLA-E and CD19 CAR ectopic expressing embryonic stem cell (ESC) line, which differentiated normally and gave rise to homogeneous CD19 CAR-NK (CD19 CAR-UiNK) cells using an organoid aggregate induction method. The CD19 CAR-UiNK were co-cultured with T cells or NK cells derived from peripheral blood mononuclear cells (PBMC) with the mismatched HLA to evaluate the immunogenicity of CD19 CAR-UiNK cells. We further assessed the therapeutic effects of CD19 CAR-UiNK cells on CD19⁺ tumor cells through in vitro cytotoxicity assays and in vivo animal models.

Results: The CD19 CAR-UiNK cells exhibited typical expression patterns of activating and inhibitory receptors, and crucial effector molecules of NK cells, similar to those of unmodified NK cells. In co-culture assays, the CD19 CAR-UiNK cells evaded allogeneic T cell response and suppressed allogeneic NK cell response. Functionally, the CD19 CAR-UiNK cells robustly secreted IFN-γ and TNF-α, and upregulated CD107a upon stimulation with Nalm-6 tumor cells. The CD19 CAR-UiNK cells effectively eliminated CD19⁺ tumor cells in vitro, including B-cell cancer cell lines and primary tumor cells from human B-cell leukemia and lymphoma. Further, the CD19 CAR-UiNK cells exhibited strong anti-tumor activity in xenograft animals.

Conclusion: We offer a strategy for deriving homogeneous and hypoimmunogenic CD19 CAR-iNK cells with robust anti-tumor effects from ESCs. Our study has significant implications for developing hypoimmunogenic CD19 CAR-NK cell therapy using human ESC as an unlimited cell source.

摘要：

背景：嵌合抗原受體（CAR）修飾的自然殺傷細(xì)胞（NK）具有不依賴mhc識(shí)別和強(qiáng)大的抗腫瘤功能等優(yōu)勢(shì)。然而，來源于人體組織的同種異體CAR-NK細(xì)胞是異質(zhì)的，容易被宿主清除。

方法：利用類器官聚集誘導(dǎo)法，制備B2M敲除、HLA-E和CD19 CAR異位表達(dá)的胚胎干細(xì)胞（ESC）系，使其正常分化并產(chǎn)生同質(zhì)的CD19 CAR- nk （CD19 CAR- uink）細(xì)胞。將CD19 CAR-UiNK細(xì)胞與HLA錯(cuò)配的外周血單個(gè)核細(xì)胞（PBMC）的T細(xì)胞或NK細(xì)胞共培養(yǎng)，評(píng)估CD19 CAR-UiNK細(xì)胞的免疫原性。該研究通過體外細(xì)胞毒性實(shí)驗(yàn)和體內(nèi)動(dòng)物模型進(jìn)一步評(píng)估了CD19 CAR-UiNK細(xì)胞對(duì)CD19+腫瘤細(xì)胞的治療作用。

結(jié)果：CD19 CAR-UiNK細(xì)胞表現(xiàn)出與未修飾NK細(xì)胞相似的活化和抑制受體以及NK細(xì)胞關(guān)鍵效應(yīng)分子的典型表達(dá)模式。在共培養(yǎng)實(shí)驗(yàn)中，CD19 CAR-UiNK細(xì)胞避開了同種異體T細(xì)胞的反應(yīng)，抑制了同種異體NK細(xì)胞的反應(yīng)。在功能上，CD19 CAR-UiNK細(xì)胞強(qiáng)烈分泌IFN-γ和TNF-α，并在Nalm-6腫瘤細(xì)胞刺激下上調(diào)CD107a。CD19 CAR-UiNK細(xì)胞在體外有效地清除了CD19+腫瘤細(xì)胞，包括b細(xì)胞癌細(xì)胞系和來自人b細(xì)胞白血病和淋巴瘤的原代腫瘤細(xì)胞。此外，CD19 CAR-UiNK細(xì)胞在異種移植動(dòng)物中表現(xiàn)出很強(qiáng)的抗腫瘤活性。

結(jié)論：該研究提供了一種從ESCs中獲得具有強(qiáng)大抗腫瘤作用的均勻性和低免疫原性CD19 CAR-iNK細(xì)胞的策略。該研究的研究對(duì)開發(fā)低免疫原性CD19 CAR-NK細(xì)胞療法具有重要意義，該療法使用人類ESC作為無限細(xì)胞來源。

該論文中，ESC、OP9細(xì)胞、表達(dá)熒光素酶的Nalm-6細(xì)胞、從三名患者的骨髓中分離出原發(fā)性人類白血病和淋巴瘤細(xì)胞的體外培養(yǎng)是使用Ausbian特級(jí)胎牛血清完成的。