Cancer Cell and Molecular Biology Branch, Division of Cancer Biology, Research Institute, National Cancer Center, 111 Jungbalsan-ro, Ilsandong-gu, Goyang, Gyeonggi-do, 410-769, Republic of Korea.
J Cancer Res Clin Oncol. 2010 Jan;136(1):143-50. doi: 10.1007/s00432-009-0645-x.
Although many efforts on revealing mechanism of the constitutive activation of NF-κB in cancer cells contributed to understanding canonical pathways, largely it remains to be determined for therapeutic approaches. Recently, we found that increased expression of transglutaminase 2 (TGase 2) appears to be responsible for constitutive activation of NF-κB in certain types of cancer cells. In previous studies, we demonstrated that TGase 2 inhibition markedly increases anti-cancer drug sensitivity in drug resistance cancer cells. Therefore, we develop safe and effective TGase 2 inhibitors for therapeutic approach.
We screened a chemical library of natural compounds using in vitro TGase 2 activity assay. The salient discovery was that glucosamine (GlcN), a known anti-inflammatory substance, inhibited the cross-linking activity of TGase 2. We tested, through a biochemical analysis including kinetics, whether the GlcN and GlcN analogs specifically inhibit TGase 2. We also determined the inhibitory mechanism using conformational change of TGase 2.
We found that the primary amine of GlcN plays a key role in TGase 2 inhibition. We also demonstrated that GlcN reversed TGase 2-mediated I-κBα polymerization in vitro. Interestingly, the metabolite of GlcN, glucosamine-6-phosphate (GlcN6P), inhibited TGase 2 activity via binding to the GTP-binding site with better efficiency than GlcN. In the native gel electrophoresis, it was clearly observed that GlcN6P binds to TGase 2 directly as an allosteric inhibitor.
We concluded that GlcN inhibits TGase 2 activity by direct contact. GlcN and its metabolite GlcN6P can down-regulate constitutive activation of NF-κB in vivo via inhibition of TGase 2.
尽管许多揭示癌细胞中 NF-κB 组成性激活机制的努力有助于理解经典途径，但在治疗方法方面仍有很大的不确定性。最近，我们发现转谷氨酰胺酶 2（TGase 2）的表达增加似乎是某些类型癌细胞中 NF-κB 组成性激活的原因。在之前的研究中，我们证明了 TGase 2 抑制显著增加了耐药癌细胞中抗癌药物的敏感性。因此，我们开发了安全有效的 TGase 2 抑制剂作为治疗方法。
我们使用体外 TGase 2 活性测定筛选了天然化合物化学文库。显著的发现是，葡萄糖胺（GlcN），一种已知的抗炎物质，抑制了 TGase 2 的交联活性。我们通过包括动力学在内的生化分析测试了 GlcN 及其 GlcN 类似物是否特异性抑制 TGase 2。我们还通过 TGase 2 的构象变化确定了抑制机制。
我们发现 GlcN 的伯胺在 TGase 2 抑制中起关键作用。我们还证明了 GlcN 在体外逆转了 TGase 2 介导的 I-κBα聚合。有趣的是，GlcN 的代谢物葡萄糖胺-6-磷酸（GlcN6P）通过与 GTP 结合位点结合抑制 TGase 2 活性的效率优于 GlcN。在天然凝胶电泳中，明显观察到 GlcN6P 作为别构抑制剂直接与 TGase 2 结合。
我们得出结论，GlcN 通过直接接触抑制 TGase 2 活性。GlcN 和其代谢物 GlcN6P 可以通过抑制 TGase 2 下调体内 NF-κB 的组成性激活。
