Update on the cloning of monoclonal anti-desmoglein antibodies from human pemphigus patients: implications for targeted therapy.
Update on the cloning of monoclonal anti-desmoglein antibodies from human pemphigus patients: implications for targeted therapy.
复制标题
人类天疱疮患者单克隆抗桥粒芯糖蛋白抗体克隆的最新进展：对靶向治疗的影响。
DOI：
10.1111/j.1365-3164.2009.00836.x
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发表时间：
2009
影响因子：
1.4
通讯作者：
Payne,AimeeS
中科院分区：
文献类型：
--
作者：
Stanley,JohnR;Ishii,Ken;Siegel,DonL;Payne,AimeeS
叶状天疱疮(PF)和寻常型天疱疮(PV)中的自身抗体分别与桥粒芯糖蛋白(DSG)1和3结合，导致角质形成细胞黏附丧失。为了鉴定这些抗体的致病性和遗传学，我们用噬菌体展示技术从患者身上分离出了单抗。利用聚合酶链式反应将外周B细胞的重链可变区和轻链可变区克隆到一个载体中，该载体产生一个噬菌体颗粒，其表面表达抗体，内部编码该抗体的cDNA.从PF或PV患者产生的噬菌体文库然后在含有DSG1或Dsg3的平板上淘洗以分离克隆。对每个克隆的DNA进行测序，以确定表达的mAb的遗传学特征。通过将来自每个独特克隆的mAb注射到正常人类皮肤器官培养中或注射到新生小鼠中，对其致病性进行测试。致病抗体会引起典型的天疱疮水泡。在PV和PF患者中，用于DSG结合抗体的重链(VH)基因都受到严格限制。PV和PF患者既有致病单抗，也有非致病单抗。抗体的免疫化学特征(包括致病性)类似于VH基因，而不是VL基因。这些单克隆致病抗体可用于筛选多肽库，以寻找阻断抗体结合的短肽。总之，抗体反应是受限的，因此，针对特定的致病抗体进行治疗可能是可行的。
Autoantibodies in pemphigus foliaceus (PF) and vulgaris (PV) bind to desmoglein (Dsg) 1 and 3, respectively, and cause loss of keratinocyte adhesion. To characterize the pathogenicity and genetics of such antibodies we have used phage display to isolate monoclonal antibodies (mAbs) from patients. PCR is used to clone the heavy and light chain variable region of the peripheral B cells into a vector that creates a phage particle with the antibody expressed on its surface and the cDNA encoding that antibody inside. The library of phage produced from a PF or PV patient are then panned on a plate containing Dsg1 or Dsg3 to isolate clones. The cDNA of each clone is sequenced to characterize the genetics of the expressed mAb. The mAb from each unique clone is tested for pathogenicity either by injecting into normal human skin organ culture or into neonatal mice. Pathogenic antibodies cause typical pemphigus blisters. In both PV and PF patients the heavy chain (VH) genes used for Dsg‐binding antibodies are severely restricted. PV and PF patients have both pathogenic and non‐pathogenic mAbs. The immunochemical characteristics of the antibodies (including pathogenicity) sort with the VH, not the VL, gene. These monoclonal pathogenic antibodies can be used to screen peptide libraries to find short peptides that block antibody binding. In summary, the antibody response is restricted and, therefore, it may be feasible to target the specific pathogenic antibodies for therapy.