同步干预2,3-DPG和PKM2在提高Bevacizumab治疗肠癌疗效中的关键作用

The limited anticancer capability of bevacizumab and its risk of promoting cancer (pro-cancer risk) included promoting metasitasis and enhancing local invasion had been recognized recently. Either upregulation of PKM2, the key enzyme of aerobic glycolysis, or increasing tumor hypoxia after administeration of bevacizumab was postulated to contribute respectively to the abovementioned pro-cancer phenomenon. Our previous studies found that bevacizumab real increased the risk of distant metastasis in mCRC patients, though it improved survival of mCRC patients; also found that GAPDH, another key enzyme of aerobic glycolysis, is a potential target for its treatment since it related to biological behavior of colorectal cancer. Furthermore, in the pilot experiments, we found that tumor became more aggressive under hypoxia condition; bevacizumab promoted liver metastasis and enhanced local invasion; colorectal cancer cell lines expressed PKM2, and its expression abviously increased after delivering bevacizumab when compared with tumor before becacizumab treatment, however, the appotosis maybe decrease after bevacizumab application; adenosine increased the concentration of 2,3-DPG in mouse RBC. We plan to improve efficacy of bevacizumab by inhibiting PKM2 since more apoptosis mediated by ROS may be induced; to overcome hypoxia via increasing of 2,3-DPG by adenosine which will prevent the pro-cancer effect of bevacizumab; simultaneous interference of 2,3-DPG and PKM2 will be more effective since the synergic effect. In addition, we plan to validate the pro-cancer risk of bevacizumab and to investage the correlation between PKM2 and its efficacy based on phase Ⅲ clinical trail (ARTIST) where bevacizuamb combined with IFL to treat mCRC in first line. The aim of study is to find the reasons of limited efficacy of bevacizumab and its solution strategy; to find efficacy predictor of bevacizumab. Finally, to resolve the problems that bevacizumab is facing in clinical practice.

Bevacizumab疗效有限性和"促癌作用"之不足刚被认识，我们推测分别与用药后有氧糖酵解关键酶PKM2上调和缺氧加重有关。基于我们前期发现bevacizumab有效治疗肠癌的同时也有促转移风险，糖酵解与肠癌生物学行为相关，也是有效的抗癌靶点；预试验发现肿瘤在缺氧环境下侵袭性增强；bevacizumab促转移和增强局部浸润力；肠癌高表达PKM2，并在bevacizumab治疗后明显升高，但细胞凋亡却有减少趋势；Adenosine显著上调红细胞内2,3-DPG（纠正缺氧）。我们拟通过抑制PKM2来增强疗效，可能机制是增加ROS介导的细胞凋亡；通过上调2,3-DPG纠正缺氧，克服"促癌作用"；同步干预因为协同作用而效果最好；在Ⅲ期临床实验资料中验证促转移风险及探讨PKM2与其疗效的关系。旨在探讨制约bevacizumab疗效的原因及其解决办法，找出疗效预测指标，为提高其临床疗效提供理论基础。

Bevacizumab是治疗转移性结直肠癌（mCRC）的重要靶向治疗药物，但是到目前为止，尚无明确的疗效预测指标。探索其疗效预测指标筛选优势人群、提高疗效是精准医疗的要求。第一，PKM2在肠癌细胞的代谢过程中起重要作用。抑制肠癌HCT-116细胞中PKM2的表达，导致细胞代谢的变化，细胞内G6PD RNA减少，而LDH RNA表达升高；细胞内GSH水平降低，ROS水平升高；葡萄糖消耗及乳酸产生均增加；细胞增殖速度下降，凋亡增强。抑制PKM2与奥沙利铂有协同作用，抑制细胞增殖、促进凋亡。第二，bevacizumab的疗效，潜在疗效预测指标及预后指标。首先，发现bevacizumab在中国mCRC患者一线、二线以及后续治疗中都是有效的。其次，治疗前血清LDH高、CA199高、左半结肠及直肠癌患者获益可能性更大。再次，发现GGT、NLR和CEA都是独立的预后指标，包含上述三个指标中0、1、2-3个指标者的生存率是显著不同的，我们建立了简单有效的预后预测模型，便于在基层医院推广。同时也发现高LDL-C/HDL-C比值、LDH和IL-8都是预后差的指标。在治疗方面，姑息性原发灶切除及局部治疗可以使部分晚期患者获益。第三，临床资料显示用过bevacizumab的患者出现肝转移的个数多、时间快。但在动物实验中，我们并未发现bevacizumab有促转移、增强局部浸润能力。第四，探索了预测bevacizumab疗效的可能机制。我们发现高LDH患者肿瘤原发灶中，乏周细胞覆盖血管（CD31+/α-SMA –）的表达率较高，而此类血管对VEGF依赖较强，对bevacizumab较为敏感。细胞实验发现，LDH表达升高，可导致肠癌细胞消耗葡萄糖及产生乳酸的量增加。而乳酸可显著抑制周细胞的迁移能力。在对于bevacizumab治疗较为敏感的左半结肠及直肠癌患者的病理标本中，也发现CD31+/α-SMA –血管的比例较高。为下一步研究提高bevacizumab疗效的新靶点提供理论基础。