背景：伴随肝功能丧失的肝转移是三阴性乳腺癌（TNBC）患者最常见的并发症之一。谱系重编程作为一种直接将功能性细胞类型从一个谱系诱导到另一个谱系而无需经过中间多能阶段的技术，有望改变细胞命运并克服原代细胞的局限性。然而，大多数重编程技术都源自人类成纤维细胞，癌细胞是否可以逆转为肝细胞仍然是个谜。方法：在这里，我们通过从两个慢病毒载体（每个表达三个因子）表达六个转录因子（HNF4A、FOXA2、FOXA3、ATF5、PROX1 和 HNF1）来简化重编程试剂的制备。然后将病毒转导到 MDA-MB-231 细胞中，生成人诱导肝细胞样细胞 (hiHeps)，并使用单细胞测序分析细胞重编程后的命运。此外，我们通过将负载有肝细胞胞外囊泡（GelMA-EV）生物墨水的明胶甲基丙烯酰水凝胶生物打印到微流控芯片上构建了肝脏芯片（LOC）模型，以评估重编程TNBC细胞在3D下的转移行为体外肝脏微环境。结果：HNF4A、FOXA2、FOXA3、ATF5、PROX1 和 HNF1A 基因的组合可以将 MDA-MB-231 肿瘤细胞重编程为人诱导肝细胞 (hiHeps)，从而限制这些细胞的转移。单细胞测序分析表明，谱系重编程后，癌基因被显着抑制，而肝脏特异性基因被激活。最后，构建的LOC模型表明，可以观察到重编程细胞的肝脏表型，并且在肝脏微环境下可以抑制嵌入癌细胞的转移。结论：我们的研究结果表明，重编程可能是一种有前途的产生肝细胞和治疗 TNBC 肝转移的方法。 LOC 模型可以模拟 3D 肝脏微环境并评估重编程 TNBC 细胞的行为。© 作者。

Background: The liver metastasis accompanied with the loss of liver function is one of the most common complications in patients with triple-negative breast cancers (TNBC). Lineage reprogramming, as a technique direct inducing the functional cell types from one lineage to another lineage without passing through an intermediate pluripotent stage, is promising in changing cell fates and overcoming the limitations of primary cells. However, most reprogramming techniques are derived from human fibroblasts, and whether cancer cells can be reversed into hepatocytes remains elusive. Methods: Herein, we simplify preparation of reprogramming reagents by expressing six transcriptional factors (HNF4A, FOXA2, FOXA3, ATF5, PROX1, and HNF1) from two lentiviral vectors, each expressing three factors. Then the virus was transduced into MDA-MB-231 cells to generated human induced hepatocyte-like cells (hiHeps) and single-cell sequencing was used to analyze the fate for the cells after reprogramming. Furthermore, we constructed a Liver-on-a-chip (LOC) model by bioprinting the Gelatin Methacryloyl hydrogel loaded with hepatocyte extracellular vesicles (GelMA-EV) bioink onto the microfluidic chip to assess the metastasis behavior of the reprogrammed TNBC cells under the 3D liver microenvironment in vitro. Results: The combination of the genes HNF4A, FOXA2, FOXA3, ATF5, PROX1 and HNF1A could reprogram MDA-MB-231 tumor cells into human-induced hepatocytes (hiHeps), limiting metastasis of these cells. Single-cell sequencing analysis showed that the oncogenes were significantly inhibited while the liver-specific genes were activated after lineage reprogramming. Finally, the constructed LOC model showed that the hepatic phenotypes of the reprogrammed cells could be observed, and the metastasis of embedded cancer cells could be inhibited under the liver microenvironment. Conclusion: Our findings demonstrate that reprogramming could be a promising method to produce hepatocytes and treat TNBC liver metastasis. And the LOC model could intimate the 3D liver microenvironment and assess the behavior of the reprogrammed TNBC cells.© The author(s).