Paul Subhojit, Chatterjee Akash, Das Kaushik, Ray Anushka, Basu Abhimanyu, Mukhopadhyay Soma, Sen Prosenjit
School of Biological Sciences, Indian Association for the Cultivation of Science, Kolkata, India.
National Institute of Biomedical Genomics, Kalyani, India.
J Biol Chem. 2025 Jan;301(1):108025. doi: 10.1016/j.jbc.2024.108025. Epub 2024 Nov 27.
The association between idiopathic venous thrombosis and occult cancer is widely recognized. However, the comprehensive understanding of how thrombin, generated during the process of thrombosis, possesses the potential to augment the malignant phenotype is still not well understood. The coagulation protease thrombin mediates its effects by cleaving protease-activated receptor 1 (PAR1), a receptor abundantly expressed on the surface of triple-negative breast cancer (TNBC) cells. While emerging evidence implicates coagulation proteases in facilitating cancer progression, the precise molecular pathways underlying thrombin-mediated induction of chemoresistance remain poorly defined. Here, we demonstrate that thrombin-induced PAR1 activation in TNBC cells promotes the development of a multidrug-resistant phenotype, mechanistically linked to the upregulation of the pro-survival protein MCL1. Genetic ablation of MCL1 sensitizes TNBC cells to cytotoxic drugs despite thrombin exposure, affirming MCL1's functional importance. Chromatin immunoprecipitation analyses reveal thrombin triggers protein kinase A-dependent phosphorylation of serine 133 residues of cAMP-responsive element-binding protein (CREB), enhancing CREB's affinity for the co-activators CBP and p300. Furthermore, thrombin treatment induces the nuclear translocation of CREB-regulated transcription coactivator 2 (CRTC2) in a calcium-dependent manner, which collectively interacts with CREB/CBP-P300. The coordinated action of these transcriptional co-activators facilitates the transcriptional induction of MCL1. We further report that PAR1 activation augments MCL1 binding to the deubiquitinase USP9X, reducing MCL1 turnover. Our pre-clinical breast cancer murine model also shows that genetic deletion of PAR1 sensitizes breast cancer cells to chemotherapeutic drugs in vivo. Collectively, these findings emphasize the thrombin-PAR1 axis as a novel driver of chemoresistance. Utilizing FDA-approved oral anticoagulants for selective blocking of thrombin action may serve as a potential therapeutic adjunct for the treatment of triple-negative breast cancer.
特发性静脉血栓形成与隐匿性癌症之间的关联已得到广泛认可。然而，对于血栓形成过程中产生的凝血酶如何具有增强恶性表型的潜力，人们仍未形成全面的理解。凝血蛋白酶凝血酶通过切割蛋白酶激活受体1（PAR1）发挥作用，PAR1是一种在三阴性乳腺癌（TNBC）细胞表面大量表达的受体。虽然越来越多的证据表明凝血蛋白酶促进癌症进展，但凝血酶介导的化疗耐药诱导的确切分子途径仍不清楚。在此，我们证明凝血酶诱导的TNBC细胞中PAR1激活促进了多药耐药表型的发展，其机制与促生存蛋白MCL1的上调有关。MCL1的基因敲除使TNBC细胞对细胞毒性药物敏感，尽管有凝血酶暴露，这证实了MCL1的功能重要性。染色质免疫沉淀分析显示，凝血酶触发蛋白激酶A依赖性的环磷酸腺苷反应元件结合蛋白（CREB）丝氨酸133残基的磷酸化，增强了CREB对共激活因子CBP和p300的亲和力。此外，凝血酶处理以钙依赖性方式诱导CREB调节的转录共激活因子2（CRTC2）的核转位，CRTC2与CREB/CBP-P300共同相互作用。这些转录共激活因子的协同作用促进了MCL1的转录诱导。我们进一步报道，PAR1激活增强了MCL1与去泛素化酶USP9X的结合，减少了MCL1的周转。我们的临床前乳腺癌小鼠模型还表明，PAR1的基因缺失使乳腺癌细胞在体内对化疗药物敏感。总体而言，这些发现强调了凝血酶-PAR1轴是化疗耐药的新驱动因素。利用FDA批准的口服抗凝剂选择性阻断凝血酶作用可能作为治疗三阴性乳腺癌的潜在治疗辅助手段。
