Abstract

Background

Enhanced anti-tumour activity is required for eradication of solid tumours by CART cells. One possibility of enhancing anti-tumour activity is by programming CART cells to express chemokine receptors that match chemokines produced either by the tumours or tumour-associated cells, thereby improving the infiltrating capacity of the CART cells. In this study, we engineered CCR2b expressing anti-Tn-MUC1 CAR T cells for the treatment of breast cancer.

Methods

Anti-Tn-MUC1-CARs were constructed using the SM3 scFv. Following lenti-MUC1 CAR retroviral transduction, efficiency of transgenic expression was assessed by flow cytometry. CCR2b expressing anti-Tn-MUC1 CAR T cells were prepared using PLV-CAR-5E5-CCR2b lentivirus. The susceptibility of MCF-7 cells to either anti-MUC1 CART or CCR2b expressing anti-MUC1 CART cell-mediated lysis was assessed using in vitro killing assays. For cytolytic analysis, CART-cells were cocultured 10:1 (effector:target) ratio with MCF-7 cells. The effects of CCR2b expressing CART cells on anti-tumour activity and infiltration were also assessed in an in vivo murine xenograft model.

Results

Activated T cells co-modified with both CCR2b and anti-MUC1-CAR had greater anti-tumour activity both in vivo and in vitro. When the effector / target cell ratio was 10, the killing rates of CART and CART-CCR2b were 56.9% and 83.9%, respectively. Tumour size was significantly smaller (P < 0.001) in the CAR-CCR2b group compared to the CAR alone group. At day 7 post-injection of CART cells, the infiltrated T cells was significantly increased (∼2 folds) in the CAR-CCR2b group compared with the CART only group.

Conclusion

Our data demonstrated that the anti-tumour activity of the CCR2b expressing anti-Tn-MUC1 CART cells is 1.5 times more potent than CART cells without CCR2b. Augmentation of tumour suppression was also demonstrated in vivo in a murine xenograft model. These pre-clinical results show translational potential to the clinic for treatment of solid breast tumours.

Legal entity responsible for the study

Guangzhou Anjie Biomedical Technology Co. Ltd.

Funding

Guangzhou Anjie Biomedical Technology Co. Ltd.

Disclosure

All authors have declared no conflicts of interest.

中文翻译：

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表达抗Tn-MUC1 CAR-T细胞的37P趋化因子受体CCR2b增强了抗乳腺癌活性

抽象的

背景

为了通过CART细胞根除实体瘤，需要增强抗肿瘤活性。增强抗肿瘤活性的一种可能性是通过对CART细胞编程以表达与肿瘤或肿瘤相关细胞产生的趋化因子相匹配的趋化因子受体，从而提高CART细胞的浸润能力。在这项研究中，我们设计了表达抗Tn-MUC1 CAR T细胞的CCR2b，用于治疗乳腺癌。

方法

使用SM3 scFv构建抗Tn-MUC1-CAR。在慢MUC1 CAR逆转录病毒转导后，通过流式细胞术评估转基因表达的效率。使用PLV-CAR-5E5-CCR2b慢病毒制备表达CCR2b的抗Tn-MUC1 CAR T细胞。使用体外杀伤试验评估了MCF-7细胞对表达抗MUC1 CART细胞介导的抗MUC1 CART或CCR2b的敏感性。为了进行细胞溶解分析，将CART细胞与MCF-7细胞以10：1（效应子：靶）的比例共培养。还在体内鼠异种移植模型中评估了表达CCR2b的CART细胞对抗肿瘤活性和浸润的影响。

结果

与CCR2b和抗MUC1-CAR共同修饰的活化T细胞在体内和体外均具有更高的抗肿瘤活性。当效应子/靶细胞比率为10时，CART和CART-CCR2b的杀死率分别为56.9％和83.9％。与单纯CAR组相比，CAR-CCR2b组的肿瘤大小明显较小（P <0.001）。注射CART细胞后第7天，与仅使用CART的组相比，CAR-CCR2b组的浸润T细胞显着增加（约2倍）。

结论

我们的数据表明，表达CCR2b的抗Tn-MUC1 CART细胞的抗肿瘤活性是没有CCR2b的CART细胞的1.5倍。还在小鼠异种移植模型中体内证明了肿瘤抑制的增强。这些临床前结果显示出可用于临床治疗实体乳腺肿瘤的转化潜力。

负责研究的法人实体

广州市安捷生物医学技术有限公司

资金

广州市安捷生物医学技术有限公司

揭露

所有作者均声明没有利益冲突。