Development of immunoregulatory therapy via CD26 activation pathway.

CD26是一种110 KDa的细胞表面糖蛋白，具有二肽基肽酶IV(DPPIV)活性，在T细胞共刺激中发挥重要作用。我们先前证明CD26与腺苷脱氨酶(ADA)直接相关，并且细胞表面表达ADA和CD26的细胞对腺苷的抑制作用更强，提示细胞表面通过CD26的ADA似乎克服了细胞外腺苷升高的抑制作用。然而，以往的研究还没有确定CD26上是否存在特定的ADA结合结构域，ADA和CD26在细胞表面的相互作用是否对T细胞激活具有直接的免疫调节作用，以及细胞表面的CD26是否涉及对腺苷抑制作用的抵抗力。我们发现CD26分子上的亮氨酸、Valine、341和gt；、丙氨酸和精氨酸是ADA结合所必需的氨基酸。当这些氨基酸被突变并导入Jurkat细胞时，得到的CD26转染体在细胞表面只表达CD26，而不表达ADA。经抗CD3+PMA刺激后，野生型和突变型CD26细胞产生IL-2的量基本相同，但突变型CD26细胞对腺苷抑制IL-2产生的敏感性明显高于野生型CD26细胞。这些数据表明，细胞表面的ADA不直接涉及T细胞的激活。相反，CD26本身并不能调节腺苷的抑制作用。只有与细胞表面CD26结合的ADA才具有功能，并能对抗细胞外腺苷升高的抑制作用。

CD26, a 110-KDa cell surface glycoprotein, exhibits dipeptidyl peptidase IV (DPPIV) enzyme activity and plays an important role in T cell costimulation. We previonsly demonstrated that CD26 is directly associated with adenosine deaminase (ADA) and that cells expressing ADA and CD26 on the surface were much more resistant to the inhibitory effect of adenosine, suggesting that ADA on the cell surface via CD26 appears to overcome the inhibitory effect of the elevated extracellular adenosine. However, previous studies have not yet determined whether there is a specific ADA binding domain on CD26, whether the interaction of ADA and CD26 on the cell surface has a direct immunoregulatory effecton T cell activation, and whether CD26 on the cell surface involves resistance to the inhibitory effect of adenosine.We found that the residues of Leucine_<340>, Valine_<341>, Alanine_<342>and Arginine_<343> on the CD26 molecule were essential amino acids for ADA binding. When these amino acids were mutated and transfected into Jurkat cells, the resultant CD26 transfectants expressed only CD26, not ADA,on the cell surface. The amount of IL-2 produced by wild-type and mutated CD26 transfectants was almost the same following stimulation with anti-CD3 plus PMA.However, the mutated CD26 transfectants were much more sensitive to the inhibitory effect of adenosine on IL-2 production than were the wild CD26 transfectants. These data suggest that ADA on the cell surface does not directlyinvolve T cell activation. Conversely, CD26 alone does not result in modulating the inhibitory effect of adenosine. Only the ADA bound to CD26 on the cell surface was functional and could counteract the inhibitory effect of elevated extracellular adenosine.