Bcl-2 interacting protein 3 (BNIP3) promotes tumor growth in breast cancer under hypoxic conditions through an autophagy-dependent pathway

Hypoxia-induced autophagy has been implicated in many cancers. Bcl-2 interacting protein 3 (BNIP3) has been associated with hypoxia, whose aberrant expression is involved in the carcinogenesis of breast cancer (BC). Here, we aim to investigate the role of hypoxia-induced autophagy and the mechanistic actions of the bioinformatically identified BNIP3 in BC. The expression pattern of BNIP3 in BC tissues and cell lines was examined using RT-qPCR and Western blot analyses. The binding affinity among BNIP3, BECN1 and BCL-2 was characterized by co-immunoprecipitation. BNIP3 expression was manipulated to assess its effects on BC cell malignant phenotypes, evaluated by cell counting kit-8, Transwell and wound healing assays, and on BC autophagy under hypoxic conditions. A BC tumor xenografts mouse model was further established to substantiate <i>in vitro</i> findings. Up-regulated expression of BNIP3 was found in BC tissues and cell lines, and BNIP3 expression was positively correlated with hypoxia exposure duration. BNIP3 knockdown restricted BC cell proliferation, invasion, and migration under hypoxic conditions. BNIP3 activated BC cell autophagy by inhibiting the binding between BCL-2 and BECN1 under hypoxic conditions. BNIP3-induced autophagy activation enhanced malignant phenotypes of BC cells, thus accelerating the tumorigenesis of BC cells <i>in vivo</i>. These data collectively supported the tumor-promoting role of BNIP3 in autophagy activation of BC under hypoxic conditions, highlighting a potential therapeutic target against BC.

缺氧诱导自噬已被证实与多种癌症的发生发展密切相关。Bcl-2相互作用蛋白3（Bcl-2 interacting protein 3, BNIP3）与缺氧过程存在关联，其异常表达参与乳腺癌（breast cancer, BC）的癌变进程。本研究旨在探讨缺氧诱导自噬的作用，以及经生物信息学筛选鉴定的BNIP3在乳腺癌中的调控机制。我们采用实时定量逆转录PCR（RT-qPCR）与蛋白质印迹法（Western blot），检测了BNIP3在乳腺癌组织及细胞系中的表达模式。通过免疫共沉淀（co-immunoprecipitation）实验，表征了BNIP3、BECN1与BCL-2三者之间的结合亲和力。通过调控BNIP3的表达水平，评估其对乳腺癌细胞恶性表型的影响——采用细胞计数试剂盒8（CCK-8）、Transwell小室实验及划痕愈合实验进行评价——同时分析其在缺氧条件下对乳腺癌细胞自噬的调控作用。进一步构建乳腺癌肿瘤异种移植小鼠模型，以验证体外实验的研究结果。实验结果显示，BNIP3在乳腺癌组织及细胞系中表达上调，且其表达水平与缺氧暴露时长呈正相关。在缺氧条件下，敲低BNIP3可抑制乳腺癌细胞的增殖、侵袭与迁移能力。缺氧环境中，BNIP3通过抑制BCL-2与BECN1的结合，激活乳腺癌细胞的自噬过程。BNIP3介导的自噬激活可增强乳腺癌细胞的恶性表型，进而在体内加速乳腺癌细胞的致瘤能力。综上，本研究证实BNIP3在缺氧条件下通过激活自噬发挥乳腺癌促瘤作用，为乳腺癌的临床治疗提供了潜在的治疗靶点。