Jain Rekha, Kulkarni Prajakta, Dhali Snigdha, Rapole Srikanth, Srivastava Sanjeeva
Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Powai, Mumbai 400076, India.
Proteomics Lab, National Centre for Cell Science, Ganeshkhind, Pune 411007, India.
J Proteomics. 2015 Jan 15;113:127-42. doi: 10.1016/j.jprot.2014.09.020. Epub 2014 Oct 5.
Glioblastoma multiforme (GBM) is one of the most devastating and dreadful WHO grade IV brain tumors associated with poor survival rate and limited therapeutics. Signal transducer and activator of transcription factor 3 (STAT3) is persistently active in several cancers, including gliomas, and STAT3 inhibitors hold great promise for treatment of glioma. LLL12, a curcumin derivative, inhibits STAT3 functions, thereby reduces growth of GBM. However, the global effects of targeting STAT3 using LLL12 have not been studied well. To shed light on this aspect, we performed quantitative proteomic analyses using differential in-gel electrophoresis (2D-DIGE) and isobaric tags for relative and absolute quantitation (iTRAQ) as well as label-free mass spectrometric analysis with 0.5μM (IC50) concentration of LLL12. Through this approach, we identified a total dataset of 1012 proteins with 1% FDR, of which 143 proteins were differentially expressed associated with various cellular functions. Results suggest that LLL12 influences central cellular metabolism and cytoskeletal proteins, in addition to its apoptosis inducing and anti-angiogenic activities, which altogether contribute to its anti-tumorigenic function. Interestingly, triose phosphate isomerase (TPI), phosphoglycerate mutase 1 (PGAM1), adaptor molecule (CRK2), protein DJ-1 (PARK7) and basic transcription factor 3 (BTF3) were found to be down-regulated and can be studied further to understand their therapeutic potential in gliomas. TPI1 and PGAM1 protein expressions were validated using immunoblot. Conclusively, our results suggest the therapeutic potential of LLL12 and it can be investigated further for a significant role in glioma treatment.
LLL12 holds great promise for therapeutic development in gliomas with constitutive expression of STAT3. This study investigated the global effect of LLL12 on the proteome of U87 glioma cells using complementary proteomic approaches, and our findings suggest that LLL12 influences central metabolism, translation, transport processes, and cytoskeleton of a cell in addition to its anti-angiogenic and apoptosis inducing functions which altogether contributes to anti-tumorigenic activity of LLL12. This study leads to the identification of several proteins which may serve as prognostic or predictive markers in GBM. We identified TPI1, PGAM1, CRK and BTF3 as potential therapeutic targets and further investigations on these candidates may facilitate therapeutic development.
多形性胶质母细胞瘤（GBM）是世界卫生组织（WHO）分级为IV级的最具破坏性和可怕的脑肿瘤之一，其生存率低且治疗方法有限。信号转导和转录激活因子3（STAT3）在包括胶质瘤在内的多种癌症中持续活跃，STAT3抑制剂在治疗胶质瘤方面具有很大的潜力。姜黄素衍生物LLL12可抑制STAT3功能，从而减少GBM的生长。然而，使用LLL12靶向STAT3的整体效果尚未得到充分研究。为了阐明这一方面，我们使用差异凝胶电泳（2D-DIGE）和相对与绝对定量的等压标签（iTRAQ）以及浓度为0.5μM（IC50）的LLL12进行无标记质谱分析，进行了定量蛋白质组学分析。通过这种方法，我们确定了一个包含1012种蛋白质的总数据集，错误发现率为1%，其中143种蛋白质与各种细胞功能相关差异表达。结果表明，LLL12除了具有诱导凋亡和抗血管生成活性外，还影响细胞的中心代谢和细胞骨架蛋白，这些共同促成了其抗肿瘤功能。有趣的是，发现磷酸丙糖异构酶（TPI）、磷酸甘油酸变位酶1（PGAM1）、衔接分子（CRK2）、蛋白质DJ-1（PARK7）和碱性转录因子3（BTF3）表达下调，可进一步研究以了解它们在胶质瘤中的治疗潜力。使用免疫印迹法验证了TPI1和PGAM1蛋白的表达。总之，我们的结果表明LLL12具有治疗潜力，可进一步研究其在胶质瘤治疗中的重要作用。
对于具有STAT3组成性表达的胶质瘤，LLL12在治疗开发方面具有很大潜力。本研究使用互补蛋白质组学方法研究了LLL12对U87胶质瘤细胞蛋白质组的整体影响，我们的研究结果表明，LLL12除了具有抗血管生成和诱导凋亡功能外，还影响细胞的中心代谢、翻译、运输过程和细胞骨架，这些共同促成了LLL12的抗肿瘤活性。本研究导致鉴定出几种可能作为GBM预后或预测标志物的蛋白质。我们将TPI1、PGAM1、CRK和BTF3鉴定为潜在的治疗靶点，对这些候选物的进一步研究可能会促进治疗开发。
